Abstract:

Disclosed are microcapsules with shells that are not animal by-products
and methods for preparing and using such microcapsules.

Claims:

1. A microcapsule, comprising: an agglomeration of primary microcapsules
and a loading substance, each individual primary microcapsule having a
primary shell, wherein the loading substance is encapsulated by the
primary shell, wherein the agglomeration is encapsulated by an outer
shell, and wherein the primary shell and the outer shell are not animal
by-products.

2-5. (canceled)

6. The microcapsule according to claim 1, wherein the primary shell or
the outer shell, or both primary and outer shells comprise a protein,
polyphosphate, polysaccharide, or mixtures thereof.

7. The microcapsule according to claim 1, wherein the primary shell or
the outer shell, or both primary and outer shells comprise a complex
coacervate.

20. The microcapsule according to claim 1, wherein the primary shell or
the outer shell, or both primary and outer shells comprise a complex
coacervate of whey protein isolate and agar, whey protein isolate and
gellan gum, whey protein isolate and gum arabic, whey protein isolate and
a caseinate, or whey protein isolate and low methoxyl pectin.

21-23. (canceled)

24. The microcapsule according to claim 1, wherein the primary shell or
the outer shell, or both primary and outer shells comprise a complex
coacervate of soy protein isolate and agar, soy protein isolate and
gellan gum, soy protein isolate and a caseinate, or pea protein isolate
and a caseinate.

25-34. (canceled)

35. The microcapsule according to claim 1, wherein the loading substance
comprises microbial oil, fungal oil, or a plant oil.

47. The microcapsule according to claim 1, wherein the loading substance
comprises an omega-3 fatty acid, an alkyl ester of an omega-3 fatty acid,
a triglyceride ester of an omega-3 fatty acid, a phytosterol ester of an
omega-3 fatty acid, arachidonic acid, and/or a mixture thereof.

51. The microcapsule according to claim 1, wherein the outer shell has an
average diameter of from about 30 μm to about 80 μm.

52-60. (canceled)

61. An emulsion, comprising a first polymer component and a loading
substance, wherein the loading substance comprises a long chain
polyunsaturated fatty acid and wherein the first polymer component is not
an animal by-product.

66. A process for preparing a microcapsule, comprising: a. providing an
emulsion comprising a first polymer component, a loading substance, and a
second polymer component, wherein the first and second polymer components
are not animal by-products; b. adjusting pH, temperature, concentration,
mixing speed, or a combination thereof to form an aqueous mixture
comprising a primary shell material, wherein the primary shell material
comprises the first and second polymer components and surrounds the
loading substance; c. further adjusting the pH, temperature,
concentration, mixing speed, or a combination thereof of the aqueous
mixture until the primary shell material forms an agglomeration; and d.
cooling the aqueous mixture to form an outer shell around the
agglomeration.

67. (canceled)

68. The process according to claim 66, further comprising step (e) spray
drying the microcapsules.

69-75. (canceled)

76. The process according to claim 66, wherein the first polymer
component the second polymer component or both the first and second
polymer components comprise a protein, polyphosphate, polysaccharide, or
mixtures thereof.

77. The process according to claim 66, wherein the first polymer
component the second polymer component or both the first and second
polymer components comprise whey protein isolate, whey protein
concentrate, soy protein isolate, soy protein concentrate, pea protein
isolate, or pea protein concentrate.

89. The process according to claim 66, wherein the primary shell or the
outer shell, or both primary and outer shells comprise a complex
coacervate.

90. The process according to claim 66, wherein the primary shell or the
outer shell, or both primary and outer shells comprise a complex
coacervate of whey protein isolate and agar, whey protein isolate and
gellan gum, whey protein isolate and gum arabic, whey protein isolate and
a caseinate, or whey protein isolate and low methoxyl pectin.

91-93. (canceled)

94. The process according to claim 66, wherein the primary shell or the
outer shell, or both primary and outer shells comprise a complex
coacervate of soy protein isolate and agar, soy protein isolate and
gellan gum, soy protein isolate and a caseinate, pea protein isolate and
a caseinate.

117. The process according to claim 66, wherein the loading substance
comprises an omega-3 fatty acid, an alkyl ester of an omega-3 fatty acid,
a triglyceride ester of an omega-3 fatty acid, a phytosterol ester of an
omega-3 fatty acid, arachidonic acid, and/or a mixture thereof.

121. The process according to claim 66, wherein cooling is at a rate of
about 1.degree. C./5 minute.

122. The process according to claim 66, wherein the mixture is cooled
until it reaches a temperature of from about 5.degree. C. to about
10.degree. C.

123-125. (canceled)

126. The process according to claim 66, wherein the outer shell has an
average diameter of from about 30 μm to about 80 μm.

127-131. (canceled)

132. A process for preparing a microcapsule, comprising: a. providing an
emulsion comprising a first polymer component, a loading substance, and a
second polymer component, wherein the first and second polymer components
are not animal by-products; b. adjusting pH, temperature, concentration,
mixing speed, or a combination thereof to form an aqueous mixture
comprising a primary shell material, wherein the primary shell material
comprises the first and second polymer components and surrounds the
loading substance; c. adjusting the pH, temperature, concentration,
mixing speed, or a combination thereof of the aqueous mixture until the
primary shell material forms an agglomeration; d. contacting the
agglomeration with a third polymer component, wherein the third polymer
component is not an animal by-product; and e. further cooling the aqueous
mixture to form an outer shell around the agglomeration.

133. (canceled)

134. The process according to claim 132, further comprising step (f)
spray drying the microcapsules.

135-141. (canceled)

142. The process according to claim 132, wherein one or more of the first
polymer component, the second polymer component, and the third polymer
component comprises a protein, polyphosphate, polysaccharide, or mixtures
thereof.

143. The process according to claim 132, wherein one or more of the first
polymer component, the second polymer component, and the third polymer
component comprises whey protein isolate, whey protein concentrate, soy
protein isolate, soy protein concentrate, pea protein isolate, or pea
protein concentrate.

155. The process according to claim 132, wherein one or more of the
primary shell, the outer shell, and the additional outer shell comprises
a complex coacervate.

156. The process according to claim 132, wherein one or more of the
primary shell, the outer shell, and the additional outer shell comprises
a complex coacervate of whey protein isolate and agar whey protein
isolate and gellan gum, whey protein isolate and gum arabic, whey protein
isolate and a caseinate, or whey protein isolate and low methoxyl pectin.

157-159. (canceled)

160. The process according to claim 132, wherein one or more of the
primary shell, the outer shell, and the additional outer shell comprises
a complex coacervate of soy protein isolate and agar, soy protein isolate
and gellan gum, soy protein isolate and a caseinate, pea protein isolate
and a caseinate.

161-166. (canceled)

167. The process according to claim 132, wherein the loading substance
comprises microbial oil, a fungal oil, or a plant oil.

179. The process according to claim 132, wherein the loading substance
comprises an omega-3 fatty acid, an alkyl ester of an omega-3 fatty acid,
a triglyceride ester of an omega-3 fatty acid, a phytosterol ester of an
omega-3 fatty acid, arachidonic acid, and/or a mixture thereof.

183. The process according to claim 132, wherein cooling is at a rate of
about 1.degree. C./5 minute.

184. The process according to claim 132, wherein the mixture is cooled
until it reaches a temperature of from about 5.degree. C. to about
10.degree. C.

185-187. (canceled)

188. The process according to claim 132, wherein the outer shell has an
average diameter of from about 30 μm to about 80 μm.

189-193. (canceled)

194. A formulation vehicle comprising a microcapsule of claim 1.

195. The formulation vehicle of claim 194, wherein the formulation
vehicle is a foodstuff, a beverage, a nutraceutical formulation, or a
pharmaceutical formulation.

196. A sachet comprising a microcapsule of claim 1.

197-201. (canceled)

202. A process for preparing a microcapsule, comprising: a. providing an
emulsion comprising a first polymer component, a loading substance, and a
second polymer component, wherein the first and second polymer components
are not animal by-products; b. adjusting pH, temperature, concentration,
mixing speed, or a combination thereof to form an aqueous mixture
comprising a primary shell material, wherein the primary shell material
comprises the first and second polymer components and surrounds the
loading substance; c. further adjusting the pH, temperature,
concentration, mixing speed, or a combination thereof of the aqueous
mixture until the primary shell material forms an agglomeration; and d.
heating the aqueous mixture to form an outer shell around the
agglomeration and to solidify the shell materials.

203. The process according to claim 66, further comprising thermally
cross-linking the shell material by heating to about 80.degree. C.

Description:

CROSS REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of priority to U.S. Provisional
Application Nos. 60/879,636, filed on Jan. 10, 2007 and 60/879,759, filed
on Jan. 10, 2007, both of which are incorporated by reference herein in
their entireties.

BACKGROUND

[0002] Many microcapsules are small particles of solids or droplets of
liquids inside a thin coating of a shell material such as beeswax,
starch, gelatin, or polyacrylic acid. They are used, for example, to
prepare liquids as free-flowing powders or compressed solids, to separate
reactive materials, to reduce toxicity, to protect against oxidation
and/or to control the rate of release of a substance such as an enzyme, a
flavor, a nutrient, a drug, etc.

[0003] In the past, research has concentrated on microcapsules where each
microcapsule had one core that contained a loading substance. However,
one of the problems with single-core microcapsules is their
susceptibility to rupture. Thus, others have tried to increase the
thickness of the microcapsule wall in order to increase the strength
and/or impermeability of such microcapsules. However, this practice can
lead to a reduction in the loading capacity of the microcapsule.

[0004] Another approach to improve microcapsules has been to create
microcapsules where each microcapsule had multiple chambers that each
contained the loading substance. For example, U.S. Pat. No. 5,780,056
discloses a "multi-core" microcapsule having gelatin as a shell material.
These microcapsules are formed by spray cooling an aqueous emulsion of
oil or carotenoid particles such that the gelatin hardens around "cores"
of the oil or carotenoid particles. Yoshida et al. (Chemical Abstract
1990:140735 or Japanese Patent Publication JP 01-148338) discloses a
complex coacervation process for the manufacture of microcapsules in
which an emulsion of gelatin and paraffin wax is added to an arabic
rubber solution and then mixed with a surfactant to form "multi-core"
microcapsules. Ijichi et al. (J. Chem. Eng. Jpn. (1997) 30(5):793-798)
micoroencapsulated large droplets of biphenyl using a complex
coacervation process to form multi-layered mirocapsules. U.S. Pat. Nos.
4,219,439 and 4,222,891 disclose "multi-nucleus" oil-containing
microcapsules having an average diameter of 3-20 μm with an oil
droplet size of 1-10 μm for use in pressure-sensitive copying papers
and heat sensitive recording papers. U.S. Pat. Nos. 6,974,592 and
6,969,530 disclose multi-nucleus oil-containing microcapsules for
delivery of various loading substances, like fish oil, to subjects.

[0005] Typically, the shell materials used to prepare such single- and
multi-core microcapsules are by-products of animals. For example,
gelatin, which has been used as a shell material for microcapsules, is
often derived from the bones, skin, and cartilage of fish, swine, and/or
cattle. While gelatin and other animal by-products are suitable
microcapsule shell materials for many purposes, they are not suitable
when one desires a microcapsule that is free of such animal by-products,
such as for religious or dietary reasons. Therefore, there is a need in
the art for microcapsules that have a high payload, are structurally
strong, and are made from shell materials that are not by-products of
animals. Disclosed herein are compositions and methods which meet these
and other needs.

SUMMARY

[0006] In accordance with the purposes of the disclosed materials,
compounds, compositions, articles, and methods, as embodied and broadly
described herein, the disclosed subject matter, in one aspect, relates to
compositions and methods for preparing and using such compositions. In a
further aspect, the disclosed subject matter relates to microcapsules
with shells that are not animal by-products and methods for preparing and
using such microcapsules. Also, in yet a further aspect, the disclosed
subject matter relates to microcapsules with shells that are prepared
from oppositely charged proteins. Methods of making and using the
disclosed microcapsules are also enclosed.

[0007] Additional advantages will be set forth in part in the description
that follows, and in part will be obvious from the description, or may be
learned by practice of the aspects described below. The advantages
described below will be realized and attained by means of the elements
and combinations particularly pointed out in the appended claims. It is
to be understood that both the foregoing general description and the
following detailed description are exemplary and explanatory only and are
not restrictive.

DETAILED DESCRIPTION

[0008] The materials, compounds, compositions, and methods described
herein may be understood more readily by reference to the following
detailed description of specific aspects of the disclosed subject matter
and the Examples included therein.

[0009] Before the present materials, compounds, compositions, and methods
are disclosed and described, it is to be understood that the aspects
described below are not limited to specific synthetic methods or specific
reagents, as such may, of course, vary. It is also to be understood that
the terminology used herein is for the purpose of describing particular
aspects only and is not intended to be limiting.

[0010] Also, throughout this specification, various publications are
referenced. The disclosures of these publications in their entireties are
hereby incorporated by reference into this application in order to more
fully describe the state of the art to which the disclosed matter
pertains. The references disclosed are also individually and specifically
incorporated by reference herein for the material contained in them that
is discussed in the sentence in which the reference is relied upon.

GENERAL DEFINITIONS

[0011] In this specification and in the claims that follow, reference will
be made to a number of terms, which shall be defined to have the
following meanings:

[0012] Throughout the specification and claims the word "comprise" and
other forms of the word, such as "comprising" and "comprises," means
including but not limited to, and is not intended to exclude, for
example, other additives, components, integers, or steps.

[0013] As used in the description and the appended claims, the singular
forms "a," "an," and "the" include plural referents unless the context
clearly dictates otherwise. Thus, for example, reference to "a compound"
includes mixtures of two or more such compounds, reference to "an omega-3
fatty acid" includes mixtures of two or more such fatty acids, reference
to "the microcapsule" includes mixtures of two or more such
microcapsules, and the like.

[0014] "Optional" or "optionally" means that the subsequently described
event or circumstance can or cannot occur, and that the description
includes instances where the event or circumstance occurs and instances
where it does not. For example the phrase "adding a loading substance, a
second polymer component, and, optionally, the composition, to the
emulsion" includes instances where the composition is added to the
emulsion and instances where the composition is not added to the
emulsion.

[0015] Ranges can be expressed herein as from "about" one particular
value, and/or to "about" another particular value. When such a range is
expressed, another aspect includes from the one particular value and/or
to the other particular value. Similarly, when values are expressed as
approximations, by use of the antecedent "about," it will be understood
that the particular value forms another aspect. It will be further
understood that the endpoints of each of the ranges are significant both
in relation to the other endpoint, and independently of the other
endpoint. It is also understood that there are a number of values
disclosed herein, and that each value is also herein disclosed as "about"
that particular value in addition to the value itself. For example, if
the value "10" is disclosed, then "about 10" is also disclosed. It is
also understood that when a value is disclosed that "less than or equal
to" the value, "greater than or equal to the value," and possible ranges
between values are also disclosed, as appropriately understood by the
skilled artisan. For example, if the value "10" is disclosed, then "less
than or equal to 10" as well as "greater than or equal to 10" is also
disclosed. It is also understood that throughout the application data are
provided in a number of different formats and that these data represent
endpoints and starting points and ranges for any combination of the data
points. For example, if a particular data point "10" and a particular
data point "15" are disclosed, it is understood that greater than,
greater than or equal to, less than, less than or equal to, and equal to
10 and 15 are considered disclosed as well as between 10 and 15. It is
also understood that each unit between two particular units are also
disclosed. For example, if 10 and 15 are disclosed, then 11, 12, 13, and
14 are also disclosed.

[0016] References in the specification and concluding claims to parts by
weight of a particular component in a composition denotes the weight
relationship between the component and any other components in the
composition for which a part by weight is expressed. Thus, in a compound
containing 2 parts by weight of component X and 5 parts by weight
component Y, X and Y are present at a weight ratio of 2:5, and are
present in such ratio regardless of whether additional components are
contained in the compound.

[0017] A weight percent (wt. %) of a component, unless specifically stated
to the contrary, is based on the total weight of the formulation or
composition in which the component is included.

[0018] "Subject," as used herein, means an individual. In one aspect, the
subject is a mammal such as a primate, and, in another aspect, the
subject is a human. The term "subject" also includes domesticated animals
(e.g., cats, dogs, etc.), livestock (e.g., cattle, horses, pigs, sheep,
goats, etc.), and laboratory animals (e.g., mouse, rabbit, rat, guinea
pig, fruit fly, etc.).

[0019] Reference herein to an "animal by-product" is meant to include
compounds and materials that are derived from, isolated from, or purified
from one or more parts of an animal's body (e.g., bones, skin, tissue,
meat, cartilage, horns, hoofs, etc.). It is also meant to include
compositions that are prepared by processing one or more animal
by-products (e.g., derivatized, functionalized, or otherwise chemically
or physically modified animal by-products). However, as used herein, an
"animal by-product" is not meant to include milk or compounds that are
derived from or isolated from animal milk, which is collected from a live
animal. Further an "animal by-product" is not meant to include eggs or
compositions derived from or isolated from eggs. The term "animal
by-product" is also not meant to include synthetic materials, or
materials derived from or isolated from plant, bacterial, fungal, or
algal sources.

[0020] The term "vegetarian" generally refers to a diet lacking meat
and/or animal by-products. It is recognized that there are various types
of vegetarian diets. For example, a vegan or total vegetarian diet
includes only foods from plants (e.g., fruits, vegetables, legumes,
grains, seeds, and nuts). A lactovegetarian diet includes food from
plants plus milk, cheese, and other dairy products. The
ovo-lactovegetarian (or lacto-ovovegetarian diet) includes food from
plants, milk, cheese, and other dairy products, and eggs. The
semi-vegetarian diet excludes red meat but includes chicken and fish,
along with foods from plants, milk, cheese, and other dairy products, and
eggs. (USDA Dietary Guidelines for Americans, 2005). Unless specifically
identified otherwise, the general term "vegetarian" as used herein
includes each of the specific types of "vegetarian" diets mentioned
above. Also, the phrase "suitable for a (particular vegetarian) diet"
means that the particular shell material or microcapsule prepared
therefrom would be acceptable for that particular vegetarian diet. For
example, a material that is obtained from eggs would be suitable for an
ovo-lactovegetarian diet (and also a semi-vegetarian diet, but not a
lactovegetarian or vegan diet). As another example, a material that is
derived from milk would be suitable for a lactovegetarian diet (and also
an ovo-lactovegetarian diet and semi-vegetarian diet, but not a vegan
diet). As yet another example, a material that is not derived from an
animal by-product, milk, or eggs, would be suitable for a vegan diet, and
for that matter a lactovegetarian, ovo-lactovegetarian, and
semi-vegetarian diet as well. In still another example, a material that
is derived from, say, fish would be suitable for a semi-vegetarian diet
(but not a lactovegetarian, ovo-lactovegetarian, or vegan diet).

[0021] Reference will now be made in detail to specific aspects of the
disclosed materials, compounds, compositions, articles, and methods,
examples of which are illustrated in the accompanying Examples.

Materials and Compositions

[0022] Disclosed herein are materials, compounds, compositions, and
components that can be used for, can be used in conjunction with, can be
used in preparation for, or are products of the disclosed methods and
compositions. These and other materials are disclosed herein, and it is
understood that when combinations, subsets, interactions, groups, etc. of
these materials are disclosed that while specific reference of each
various individual and collective combinations and permutation of these
compounds may not be explicitly disclosed, each is specifically
contemplated and described herein. For example, if a compound is
disclosed and a number of modifications that can be made to a number of
components or residues of the compound are discussed, each and every
combination and permutation that are possible are specifically
contemplated unless specifically indicated to the contrary. Thus, if a
class of components A, B, and C are disclosed as well as a class of
components D, E, and F and an example of a combination composition A-D is
disclosed, then even if each is not individually recited, each is
individually and collectively contemplated. Thus, in this example, each
of the combinations A-E, A-F, B-D, B-E, B-F, C-D, C-E, and C-F are
specifically contemplated and should be considered disclosed from
disclosure of A, B, and C; D, E, and F; and the example combination A-D.
Likewise, any subset or combination of these is also specifically
contemplated and disclosed. Thus, for example, the sub-group of A-E, B-F,
and C-E are specifically contemplated and should be considered disclosed
from disclosure of A, B, and C; D, E, and F; and the example combination
A-D. This concept applies to all aspects of this disclosure including,
but not limited to, steps in methods of making and using the disclosed
compositions. Thus, if there are a variety of additional steps that can
be performed it is understood that each of these additional steps can be
performed with any specific aspect or combination of aspects of the
disclosed methods, and that each such combination is specifically
contemplated and should be considered disclosed.

Microcapsules

[0023] In certain examples, disclosed herein are microcapsules that
comprise an agglomeration of primary microcapsules and a loading
substance, each individual primary microcapsule having a primary shell,
wherein the loading substance is encapsulated by the primary shell and
the agglomeration is encapsulated by an outer shell, and wherein the
primary shell and the outer shell are not animal by-products. These
microcapsules are referred to herein as "multicore microcapsules." Also
disclosed are "single-core" microcapsules that comprise a core, wherein
the core comprises a loading substance, a primary shell surrounding the
core, and an outer shell surrounding the primary shell, wherein the
primary shell and the outer shell are not animal by-products. Unless
stated otherwise, the term "microcapsule" is used herein to refer to
multicore, single-core, or a mixture of multicore and single-core
microcapsules. In these microcapsules (and others disclosed herein) the
primary and outer shells comprise a non-animal by product, as is defined
herein. Still further, disclosed are microcapsules comprising a loading
substance and a polymer component, wherein the loading substance is
surrounded by the polymer component, wherein the loading substance
comprises a long chain polyunsaturated fatty acid, and wherein the
polymer component is not an animal by-product.

[0024] Also, disclosed herein are microcapsules that comprise an
agglomeration of primary microcapsules and a loading substance, each
individual primary microcapsule having a primary shell, wherein the
loading substance is encapsulated by the primary shell and the
agglomeration is encapsulated by an outer shell, and wherein the primary
shell and the outer shell are suitable for one or more of a vegan diet
(e.g., the shells are not obtained from animal by-products, milk, or
eggs), a lactovegetarian diet (e.g., the shells are not obtained from
animal-by products or eggs, but can be obtained from milk), or a
ovo-lactovegetarian diet (e.g., the shells are not obtained from
animal-by products, but may be obtained from milk or eggs). In other
examples, the primary and outer shells are suitable for a semi-vegetarian
diet (e.g., the shells are obtained from fish).

[0025] Further, disclosed herein are microcapsules that comprise shells
made from two oppositely charged proteins. That is, in the disclosed
microcapsules the shell materials can be complex coacervates formed from
two or more oppositely charged polymers. In certain particular examples,
the oppositely charges polymers are both proteins. For example, disclosed
herein are microcapsules where the shell materials (primary and/or outer
shells) are complex coacervates form from a positively charged protein
(such as whey, pea, or soy protein isolates or concentrates) and a
negatively charged protein (such as caseinate) instead of a polyanionic
polymer like gum aracaia.

[0026] The term "residue" as used herein refers to the moiety that is the
resulting product of the specified chemical species in a particular
reaction scheme or subsequent formulation or chemical product, regardless
of whether the moiety is actually obtained from the specified chemical
species. For example, an "amino acid residue" refers to the moiety which
results when an amino acid participates in a particular reaction (e.g.,
the residue can be the product of an amino acid undergoing a
transglutaminase catalyzed crosslinking reaction with another amino
acid). In this case, the amino acid residue is "derived" from the amino
acid. It is understood that this moiety can be obtained by a reaction
with a species other than the specified amino acid, for example, by a
reaction with a protein or peptide containing the amino acid, and the
like. This concept applies to other chemical species disclosed herein,
such as protein, saccharides like chitosan, lactose, and sucrose, and
waxes. Thus, when such species undergo particular reactions or treatment
(e.g., acid/base reactions, crosslinking reactions with other chemical
species, and functional group transformations), they are referred to
herein as a residue of the corresponding chemical species.

[0027] It is also contemplated that one or more additional shell layers
can be placed on the outer shell of the microcapsules. The techniques
described in International Publication No. WO 2004/041251 A1, which is
incorporated by reference in its entirety, can be used to add additional
shell layers to the microcapsules. It is understood, however, that the
additional shell materials are not animal by-products.

[0028] Shell Materials

[0029] A number of different polymers can be used to produce the shell
layers of the disclosed single-core and multicore microcapsules. For
example, the primary shell and/or outer shell material of the disclosed
microcapsules can comprise a protein, polyphosphate, polysaccharide, or
mixtures thereof, which are not animal by-products. The disclosed
microcapsules can contain shells that are coacervates of two oppositely
charged polymers. For example, a polymer that is cationic or can be made
cationic by adjustments in pH can be combined with a polymer that is
anionic or can be made anionic by adjustments in pH to form a coacervate
shell. In certain examples, the cationic polymers and anionic polymers
are both proteins.

[0030] A particularly suitable shell material that is not an animal
by-product as defined herein is whey protein. Whey protein typically
comes in two major forms: isolate and concentrate. Unless specifically
stated to the contrary the terms whey protein isolate and whey protein
concentrate are included in the meaning of the term "whey protein." Whey
protein concentrates contain fat, lactose, carbohydrates, and bioactive
compounds. Whey protein isolates are processed to remove the fat,
lactose, and carbohydrates, yet are usually lower in bioactive compounds
as well. Generally speaking, whey protein isolate (WPI) is a collection
of globular proteins that is isolated from whey, which is typically a
by-product of cheese manufactured from bovine milk. In this sense, whey
protein (isolates and concentrates) are suitable for lactovegetarian,
ovo-lactovegtarian, and semi-vegetarian diets. WPI is a mixture of
β-lactoglobulin (about 65%), α-lactoglobulin (about 25%), and
serum albumin (about 8%), which are soluble in their native forms,
independent of pH. WPI can be nearly 90% protein by weight. WPI can also
include trace amounts of immunoglobulins IgG, IgA and IgM,
glycomacropeptides, lactoferrin, lactoperoxidase, and/or lysozyme. WPI
can be obtained from commercial sources such as NZMP ALACEN 895® from
Nealanders International Inc. (Rocky River, Ohio).

[0031] Another suitable shell material that is not an animal by-product as
defined herein is soy protein, which includes soy protein concentrates
and isolates. Soy protein isolates (SPI) is a highly refined or purified
form of soy protein with a minimum protein content of about 90% on a dry
basis. It is made from defatted soy flour, which has had most of the
non-protein components, fats, and carbohydrates removed. It is typically
used as a health food because it is a complete vegetable containing all
the essential amino acids for growth. Also, it has a very low fat content
when compared to animal sources of protein, such as meat or milk. SPI can
be obtained from commercial sources such as PRO FAM 781® from ADM
Protein Specialties Division (Decatur, Ill.). Soy protein (isolates and
concentrates) can be suitable for vegan, lactovegetarian,
ovo-lactovegetarain, and semi-vegetarian diets.

[0032] Still another suitable shell material that is not an animal
by-product as defined herein is pea protein, which includes pea protein
concentrates and isolates. Pea protein can be obtained from a variety of
species of pea. Pea protein isolates and concentrates can be obtained
from commercial sources such as Roquette America, Inc., (Keokuk, Iowa)
and Kirkman (Lake Oswego, Oreg.). Pea protein can be suitable for vegan,
lactovegetarian, ovo-lactovegetarain, and semi-vegetarian diets.

[0033] Caseins are further examples of suitable shell materials that are
not animal by-products. Caseins account for about 80% of the total
protein in bovine milk, while whey proteins account for the remaining
approximately 20%. Caseins are produced by precipitation with either acid
at about pH 4.6 or rennet enzyme and the subsequent drying of the
precipitate. Caseins are not typically coagulated by heat, do not
denature, and are relatively hydrophobic. Caseinates are solubilized
forms of casein produced by reaction with an alkaline substance. Common
caseinates include: sodium caseinate, calcium caseinate, potassium
caseinate, and ammonium caseinate. "Caseinates" is used herein to
generally refer to these and other caseinates. Sodium caseinate is highly
soluble and is used as an emulsifier in coffee whiteners, cottage cheese,
cream liqueurs, yogurt, processed cheeses, and some meat products.
Caseins and caseinates are commercially available and are suitable for
lactovegetarian, ovo-lactovegetarain, and semi-vegetarian diets.

[0034] Egg white protein, which is a suitable shell material that is not
an animal by-product as defined herein, also called albumin, is soluble
in water, insoluble in alcohol or ether, and is used in food systems for
foaming and gelation. On heating an aqueous solution of egg white protein
to about 75° C., it becomes coagulated. Egg white protein can be
suitable for ovo-lactovegetarain and semi-vegetarian diets.

[0035] Cereal prolamine proteins are still further examples of shell
materials that are not animal by-products as defined herein. Cereal
prolamine proteins are insoluble in water and anhydrous alcohol, and
soluble in a mixture of the two. Zein, found in maize, is one of the most
well understood plant proteins. It is clear, odorless, tasteless, hard,
water-insoluble and edible, used as a coating for candy, nuts, fruit,
pills, and other encapsulated foods and drugs, labeled as "confectioner's
glaze" or as "vegetable protein", a very good water barrier, offering
extended shelf-life, particularly under high-humidity and high-heat
condition. Cereal prolamine proteins can be suitable for vegan,
lactovegetarian, ovo-lactovegetarain, and semi-vegetarian diets.

[0036] Still another suitable shell material that is not an animal
by-product as defined herein is agar. Agar is a polymer made up of
subunits of galactose. It is a component of algae cell walls. It is a
vegetarian substitute for gelatin and is even firmer and stronger than
gelatin. Agar gels around 32-40° C. and remains solid up to about
85° C. Its major use is as a culture medium for microbiological
work but another use is as a laxative. Agar performs well during complex
coacervates as a polyanion. Agar can be obtained from commercial sources
such as AGAR RS-100® from TIC Gums (Belcamp, Md.). Agar can be
suitable for vegan, lactovegetarian, ovo-lactovegetarain, and
semi-vegetarian diets.

[0037] Gellan gum is another suitable shell material that is not an animal
by-product as defined herein and can be used in the compositions and
methods disclosed herein. Gellan gum is a vegetarian gelatin substitute
and is a polysaccharide produced by the bacterium Sphingomonas elodea,
which is soluble in water. It is used primarily as an alternative to agar
as a gelling agent in microbiological culture. In certain applications,
gellan gum can be more desirable than agar because it has better visual
clarity and strength and it is able to withstand temperatures of about
120° C.; thus, it is safe during spray-drying processes. Also, one
needs only approximately half the amount of gellan gum as agar to reach
an equivalent gel strength, though the exact texture and quality depends
on the concentration of divalent cations present. As a food additive,
gellan gum is used as a thicker, emulsifier and stabiliser. Gellan gum
can be obtained from commercial sources such as from KELCOGEL F® from
C.P. Kelco (San Diego, Calif.). Gellan gum can be suitable for vegan,
lactovegetarian, ovo-lactovegetarain, and semi-vegetarian diets.

[0038] Gum arabic is yet another suitable shell material that is not an
animal by-product as disclosed herein and can be used in the compositions
and methods disclosed herein. Gum arabic is a substance is taken from two
sub-Saharan species of the acacia tree, Acacia senegal and Acacia seyal.
It is used primarily in the food industry as a stabilizer, but has had
more varied uses in the past, including viscosity control in inks. Its E
number is E-414. Gum arabic is a complex mixture of saccharides and
glycoproteins, and it is edible. It is an ingredient in soft drink
syrups, "hard" gummy candies like gumdrops, marshmallows, and most
notably, chewing gums. For artists it is the traditional binder used in
watercolor paint, and was used in photography for gum printing.
Pharmaceuticals and cosmetics also use gum arabic. Gum Arabic can be
obtained from commercial sources such as TIC gums (Belcamp, Md.).

[0039] Xanthan gum is still another suitable shell material that is not an
animal by-product as defined herein. Xanthan gum is a natural gum
polysaccharide as a food additive and rheology modifier. It is produced
by a biotechnological process involving fermentation of glucose or
sucrose by Xanthomonas campestris. One of the properties of xanthan gum
is its capability of producing a large increase in the viscosity by
adding a very small quantity of gum (e.g., on the order of one percent).
In most foods, it is used at 0.5% or as low as 0.05%. The viscosity of
xanthan gum solutions decreases with higher shear rates. Like other gums
it is very stable under a wide range of temperatures and pH. Xanthan gum
is commercially available. Xanthan gum can be suitable for vegan,
lactovegetarian, ovo-lactovegetarain, and semi-vegetarian diets.

[0040] Pectin is yet another suitable shell material that is not an animal
by-product as defined herein. Pectin is a grouping of acid structural
polysaccharides found in fruit and vegetables and is prepared mainly from
citrus peel waste and apple pomace. Pectin can be used as a replacement
for polyphosphate, which has been used as a shell material, because it is
abundant and relatively inexpensive. Amidated pectin is suitable gelatin
replacement or supplement because it also has amine portions on its
structure that can be crosslinked by mechanisms similar to that used for
gelatin. This allows for a quicker development cycle compared to having
to develop a new technology or modification of current technology.
Low-methoxyl-pectin is also a suitable shell material. Pectin and
low-methoxyl pectin can be suitable for vegan, lactovegetarian,
ovo-lactovegetarain, and semi-vegetarian diets.

[0042] The shell material can be a two-component system made from a
mixture of different types of polymer components, and where a composition
has been added to the system to improve impermeability. In other
examples, the shell material can be a complex coacervate between two or
more polymer components (e.g., whey or soy protein isolate and agar).
Component A can be whey or soy protein isolate, although other polymers
like those mentioned above for the shell materials are also contemplated
as component A. Component B can be agar, gellan gum, pectin, low methoxyl
pectin, gum arabic, alginate, chitosan, carrageenan,
carboxymethyl-cellulose or a mixture thereof. Again other polymers like
those disclosed above for the shell materials are also contemplated as
component B. The molar ratio of component A:component B that is used
depends on the type of components but is typically from about 1:5 to
about 15:1. For example, when whey or soy protein isolate and agar are
used as components A and B respectively, the molar ratio of component
A:component B can be about 8:1 to about 12:1; when whey or soy protein
isolate and gellan gum are used as components A and B respectively, the
molar ratio of component A:component B can be about 2:1 to about 1:2; and
when whey or soy protein isolate and low methoxyl pectin are used as
components A and B respectively, the molar ratio of component A:component
B can be about 3:1 to about 5:1. In many of the disclosed microcapsules
the primary shell and/or outer shell can comprise a complex coacervate.
For example, the primary shell and/or outer shell can comprise a complex
coacervate of whey, pea, or soy protein isolate and agar and/or gellan
gum. In other examples, the primary and/or outer shell can comprise a
complex coacervate of whey, pea, or soy protein isolate and a caseinate
(e.g., sodium, calcium, potassium, or ammonium caseinate).

[0043] In particular examples, using WPI, PPI, or SPI and agar to prepare
primary microcapsules, and having gellan gum deposit on the surface of
the primary microcapsules to form an outer shell, results in stable
non-gelatin or vegetarian microcapsules without the need for any
transglutaminase crosslinking. Likewise, using WPI, PPI, or SPI and gum
arabic or caseinate can result in compact microcapsules with long
induction periods. Further, the use of such shell materials can lower
costs significantly since transglutaminase is expensive and requires long
production time.

[0044] In the disclosed microcapsules the outer shell can have an average
diameter of from about 1 μm to about 2,000 μm, from about 20 μm
to about 1,000 μm, or from about 30 μm to about 80 μm. In
further examples, the average diameter of the outer shell can be about 1,
10, 20, 30, 40, 50, 60, 70, 80, 90, 200, 300, 400, 500, 600, 700, 800,
900, 1000, 1200, 1300, 1400, 1500, 1600, 1700, 1800, 1900, or 2000 μm,
where any of the stated values can form an upper or lower endpoint when
appropriate.

[0045] The primary shells of the disclosed microcapsules can have an
average diameter of from about 40 nm to about 10 μm or from about 0.1
μm to about 5 μm. In further examples, the average diameter of the
primary shell can be about 40 nm, 50 nm, 60 nm, 70 nm, 80 nm, 90 nm, 100
nm, 200 nm, 300 nm, 400 nm, 500 nm, 600 nm, 700 nm, 800 nm, 900 nm, 1000
nm, 2 μM, 3 μm, 4 μm, 5 μm, 6 μm, 7 μm, 8 μm, 9
μm, 10 μm, where any of the stated values can form an upper or
lower endpoint when appropriate. Particle size can be measured using any
typical equipment known in the art, for example, a COULTER® LS230
Particle Size Analyzer, Miami, Fla., USA.

[0046] Loading Substances

[0047] In the disclosed microcapsules, the loading substance can be any
substance that one desires to be microencapsulated (e.g., a substance
that one desired to be delivered to a subject). In many examples, a
suitable loading substance is not entirely soluble in an aqueous mixture.
The loading substance can be a solid, a hydrophobic liquid, or a mixture
of a solid and a hydrophobic liquid. In many of the examples herein, the
loading substance can comprise a long chain polyunsaturated fatty acid,
specific examples of which are included below. Further, the loading
substance can comprise a biologically active substance, a nutrient such
as a nutritional supplement, a flavoring substance, a polyunsaturated
fatty acid like an omega-3 fatty acid, a vitamin, a mineral, a
carbohydrate, a steroid, a trace element, and/or a protein, and the like
including mixtures and combinations thereof. In other examples, the
loading substance can comprise microbial oil, algal oil (e.g., oil from a
dinoflagellate such as Crypthecodinium cohnii), fungal oil (e.g., oil
from Thraustochytrium, Schizochytrium, or a mixture thereof), and/or
plant oil (e.g., flax, vegetables), including mixtures and combinations
thereof. In other examples, the loading substance can be a pharmaceutical
composition (e.g., a drug and/or an enzyme) or a flavor. The loading
substance can also be a hydrophobic liquid, such as grease, oil or a
mixture thereof. Typical oils can be fish oils, vegetable oils (e.g.,
canola, olive, corn, rapeseed), mineral oils, derivatives thereof or
mixtures thereof. The loading substance can comprise a purified or
partially purified oily substance such as a fatty acid, a triglyceride,
or a mixture thereof.

[0049] Many of the microbial, algal, fungal, plant, and marine oils
disclosed herein contain omega-3 fatty acids. As such, certain delivery
devices disclosed herein can contain a loading substance that comprises
an omega-3 fatty acid, an alkyl ester of an omega-3 fatty acid, a
triglyceride ester of an omega-3 fatty acid, a phytosterol ester of an
omega-3 fatty acid, and/or mixtures and combinations thereof. An omega-3
fatty acid is an unsaturated fatty acid that contains as its terminus
CH3--CH2--CH═CH--. Generally, an omega-3 fatty acid has the
following formula:

##STR00001##

wherein R1 is a C3-C40 alkyl or alkenyl group comprising
at least one double bond and R2 is H or alkyl group. The term
"alkane" or "alkyl" as used herein is a saturated hydrocarbon group
(e.g., methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, s-butyl,
t-butyl, n-pentyl, isopentyl, s-pentyl, neopentyl, hexyl, heptyl, octyl,
nonyl, decyl, dodecyl, tetradecyl, hexadecyl, eicosyl, tetracosyl, and
the like). The term "alkene" or "alkenyl" as used herein is a hydrocarbon
group containing at least one carbon-carbon double bond. Asymmetric
structures such as (AB)C═C(CD) are intended to include both the E and
Z isomers (cis and trans). In a further example, R1 can be a
C5-C38, C6-C36, C8-C34, C10-C32)
C12-C30, C14-C28, C16-C26, or
C18-C24 alkenyl group. In yet another example, the alkenyl
group of R1 can have from 2 to 6, from 3 to 6, from 4 to 6, or from
5 to 6 double bonds. Still further, the alkenyl group of can have from 1,
2, 3, 4, 5, or 6 double bonds, where any of the stated values can form an
upper or lower endpoint as appropriate.

[0050] Specific examples of omega-3 fatty acids that are suitable loading
substances that can be used in the disclosed delivery devices include,
but are not limited to, α-linolenic acid (18:3ω3),
octadecatetraenoic acid (18:4ω3), eicosapentaenoic acid
(20:5ω3) (EPA), eicosatetraenoic acid (20:4ω3),
henicosapentaenoic acid (21:5ω3), docosahexaenoic acid
(22:6ω3) (DHA), docosapentaenoic acid (22:5ω3) (DPA),
including derivatives and mixtures thereof. Many types of fatty acid
derivatives are well known to one skilled in the art. Examples of
suitable derivatives are esters, such as phytosterol esters, furanoid
esters, branched or unbranched C1-C30 alkyl esters, branched or
unbranchcd C2-C30 alkenyl esters or branched or unbranched
C3-C30 cycloalkyl esters, in particular phytosterol esters and
C1-C6 alkyl esters. In a further example, the loading substance
can be a phytosterol ester of docosahexaenoic acid and/or
eicosapentaenoic acid, a C1-C6 alkyl ester of docosahexaenoic
acid and/or eicosapentaenoic acid, a triglyceride ester of
docosahexaenoic acid and/or eicosapentaenoic acid, and/or a mixture
thereof.

[0051] Other examples of suitable loading substances that can be present
in the disclosed delivery devices comprise at least 4, at least 6, at
least 8, at least 10, at least 12, at least 14, at least 16, at least 18,
or at least 20 carbon atoms. In some other examples, the loading
substance can contain about 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19,
20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37,
38, 39, 40, 41, 42, 43, 44, or 45 carbon atoms, where any of the stated
values can form an upper or lower endpoint when appropriate. In still
other examples, the loading substance can comprise a mixture of fatty
acids (including derivatives thereof) having a range of carbon atoms. For
example, the loading substance can comprise from about 8 to about 40,
from about 10 to about 38, from about 12 to about 36, from about 14 to
about 34, from about 16 to about 32, from about 18 to about 30, or from
about 20 to about 28 carbon atoms.

[0052] Some further examples of loading substances are those that contain
at least one unsaturated bond (i.e., a carbon-carbon double or triple
bond). For example, the loading substance can contain at least 2, at
least 3, at least 4, at least 5, at least 6, at least 7, or at least 8
carbon-carbon double bonds, triple bonds, or any combination thereof. In
another example, the loading substance can comprise 1, 2, 3, 4, 5, 6, 7,
or 8 unsaturated bonds, where any of the stated values can form an upper
or lower endpoint as appropriate.

[0053] Some specific examples of loading substances, which are unsaturated
fatty acids, are shown in the following tables. Derivatives of these
fatty acids are also suitable and are thus contemplated herein.

[0055] In the above paragraph (and throughout) the compounds are
identified by referring first to the "n-x family," where x is the
position in the fatty acid where the first double bond begins. The
numbering scheme begins at the terminal end of the fatty acid, where, for
example, the terminal CH3 group is designated position 1. In this
sense, the n-3 family would be an omega-3 fatty acid, as described above.
The next number identifies the total number of carbon atoms in the fatty
acid. The third number, which is after the colon, designates the total
number of double bonds in the fatty acid. So, for example, in the n-1
family, 16:3, refers to a 16 carbon long fatty acid with 3 double bonds,
each separated by a methylene, wherein the first double bond begins at
position 1, i.e., the terminal end of the fatty acid. In another example,
in the n-6 family, 18:3, refers to an 18 carbon long fatty acid with 3
methylene separated double bonds beginning at position 6, i.e., the sixth
carbon from the terminal end of the fatty acid, and so forth.

[0056] Further examples of loading substances that contain at least one
pair of methylene interrupted unsaturated bonds are shown in Table 2.

[0057] Specific examples of suitable loading substances that contain
conjugated unsaturated bonds include, but are not limited to, those in
Table 3. By "conjugated unsaturated bond" is meant that at least one pair
of carbon-carbon double and/or triple bonds are bonded together, without
a methylene (CH2) group between them (e.g.,
--CH═CH--CH═CH--).

[0058] In the above examples of suitable loading substances, derivatives
of the disclosed loading substances can also be used. By "derivatives" is
meant the ester of a fatty acid (e.g., methyl and ethyl esters), salts of
the fatty acids (e.g., sodium and potassium salts), and triglycerides,
diglycerides, and monoglycerides, sterol esters, antioxidant-oil
conjugates (e.g., ascorbyl palmitate), and naturally derivatives such as
furanoid fatty acid derivatives.

[0059] The loading substances disclosed herein can also be crude oils,
semi-refined (also called alkaline refined), or refined oils from such
sources disclosed herein. Still further, the disclosed compositions and
methods can use oils comprising re-esterified triglycerides.

[0060] It is contemplated herein that one or more of the disclosed loading
substances can be used. For example the disclosed delivery devices can
contain two or more different loading substances. Further, the loading
substance can be present in an amount of from about 1% to about 50% by
weight of a microcapsule. In specific examples, the loading substance can
be present in an amount of from about 1% to about 40%, from about 1% to
about 30%, from about 1% to about 20%, from about 1% to about 15%, or
from about 1% to about 10% by weight of a microcapsule.

[0061] In one example, the loading substance is not a fatty acid
conjugate. A fatty acid conjugate is a fatty acid that has been coupled
to (e.g., bonded to) another chemical moiety, such as a metal (e.g.,
chromium) or cofactor (CoQ10). In other examples, the loading
substance is not oil with a low interfacial tension (IT) (i.e., having an
interfacial tension of less than about 15 dynes/cm). In other examples,
the loading substance is such a fatty acid conjugate or low IT oil.

[0062] In one example, the loading substances can be or can contain an
antioxidant. Suitable examples of antioxidants include, but are not
limited to, a phenolic compound, a plant extract, or a sulfur-containing
compound. In certain examples disclosed herein the antioxidant can be
ascorbic acid or a salt thereof, e.g., sodium ascorbate. In other
examples, the antioxidant can be citric acid or a salt thereof. In still
other examples, the antioxidant can be vitamin E, CoQ10, lutein,
zeaxanthan, carotene (e.g., beta-carotene) tocopherols, lipid soluble
derivatives of more polar antioxidants such as ascobyl fatty acid esters
(e.g., ascobyl palmitate), plant extracts (e.g., rosemary, sage and
oregano oils), algal extracts, and synthetic antioxidants (e.g., BHT,
TBHQ, ethoxyquin, alkyl gallates, hydroquinones, tocotrienols), or
mixtures thereof.

[0063] The disclosed loading substance can also be or contain other
nutrient(s) such as vitamins other trace elements (e.g., zinc), minerals,
and the like. Further, the loading substances can comprise other
components such as preservatives, antimicrobials, anti-oxidants,
chelating agents, thickeners, flavorings, diluents, emulsifiers,
dispersing aids, or binders, including any mixture thereof.

[0064] In addition, the loading substance can have a low interfacial
tension. For example, a suitable loading substance can have an
interfacial tension of less than about 20, less than about 15, less than
about 11, less than about 9, less than about 7, or less than about 5
dynes/cm. In other examples, the loading substance can have an
interfacial tension of from about 0.1 to about 20, from about 1 to about
15, from about 2 to about 9, from about 3 to about 9, from about 4 to
about 9, from about 5 to about 9, or from about 2 to about 7 dynes/cm. In
still further examples, the loading substance can have an interfacial
tension of about 0.1, 0.5, 1.0, 1.5, 2.0, 2.5, 3.0, 3.5, 4.0, 4.5, 5.0,
5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, 9.5, 10.0, 10.5, 11.0, 11.5,
12.0, 12.5, 13.0, 13.5, 14.0, 14.5, 15.0, 15.5, 16.0, 16.5, 17.0, 17.5,
18.0, 18.5, 19.0, 19.5, or 20.0, where any of the stated values can form
an upper or lower endpoint when appropriate. In particular examples, the
loading substance can be an algal oil with an interfacial tension of
about 0.5, 0.6, 0.7, 0.8, 0.9, or 1.0 dynes/cm. The loading substance can
also be a fungal oil with an interfacial tension of about 3.0, 3.1, 3.2,
3.3, or 3.4 dynes/cm.

[0065] The interfacial tension of a loading substance can be determined by
methods known in the art. For example, the interfacial tension from a
loading substance to a standard gelatin solution or from a loading
substance to distilled water can be determined with a Fisher Surface
Tensiomat. Generally, a standard gelatin solution or distilled water can
be poured into a sample vessel, which is placed on the sample table of a
tensiomat. The loading substance can then be added to the sample vessel.
The sample can be raised so that the ring of the tensiomat is immersed in
the loading substance. The interfacial tension is the measure of downward
force on the ring as it passes through the interface of the loading
substance and standard gelatin solution or the interface of the loading
substance and distilled water, depending on whichever experimental setup
is being used.

[0067] Further, the payloads of loading substances in the disclosed
microcapsules can be from about 20% to about 90%, about 50% to about 70%
by weight, or about 60% by weight of the microcapsule. In other examples,
the disclosed microcapsules can contain about 20, 25, 30, 35, 40, 45, 50,
55, 60, 65, 70, 75, 80, 85, or 90% by weight of the microcapsule, where
any of the stated values can form an upper or lower endpoint when
appropriate.

SPECIFIC EXAMPLES

[0068] Specific examples of microcapsules that contain any of the shell
materials and any of the loading substances are disclosed herein. Some
specific examples include, but are not limited to, microcapsules where
the shell materials are complex coacervates, e.g., coacervates of whey
protein isolates and agar, gellan gum, gum arabic, caseinate, and/or low
methoxyl pectin. In another example, the microcapsules can have shell
materials that are complex coacervates of soy protein isolates and agar,
gellan gum, gum arabic, caseinate, and/or low methoxyl pectin. In still
another example, the microcapsules can have shell materials that are
complex coacervates of pea protein isolate and agar, gellan gum, gum
arabic, caseinate, and/or low methoxyl pectin. Loading substances that
can be used can, in many instances, include marine oils (e.g., fish oils
and algal oils). Loading substances that comprise omega-3 fatty acids
such as EPA and DHA can also be desirable. Further, derivatives of
omega-3 fatty acids, such as mono-, di-, and triglycerides, alkyl esters,
sterol esters, antioxidant esters (e.g., ascorbyl and citryl esters), and
furanoid esters, can also be suitable loading substances.

[0069] Some particularly suitable microcapsules include microcapsules
containing fish oils. Examples of such fish oils include, but are not
limited to, sardine, anchovy, bonito, and/or tuna oil. Fish oils can also
be referred to herein by the approximate ratio of EPA and DHA, or
derivatives thereof, found in the oil. For example, 18:12 oils generally
comprise a ratio of EPA to DHA (or their triglyceride esters for example)
of about 18:12. Likewise, 5:25 oils generally comprise a ratio of EPA to
DHA of about 5:25. Such microcapsules can be Generally Regarded as Safe
(GRAS), kosher, and/or Halal. Further, such microcapsules can contain
algal oils comprising omega-3 fatty acids. In this case, the
microcapsules can be regarded as organic, vegetarian, and/or vegan,
depending on the particular shell material and the particular standards
for classifying such materials. Also, such microcapsules can have at
least about 130 mg of DHA or at least about 150 mg of EPA and DHA per
gram of powder. Further, antioxidants such as ascorbic acid, citric acid,
and/or phosphoric acid (or salts thereof) can be present in such
microcapsules.

Emulsions

[0070] Also disclosed herein are emulsions that comprise a first polymer
component and a loading substance, wherein the loading substance
comprises a long chain polyunsaturated fatty acid and wherein the first
polymer component is not an animal by-product. Any of the loading
substances disclosed herein can be used. For example the loading
substance can comprise an omega-3 fatty acid. The loading substance can
comprise a marine oil. The loading substance can comprise a fish oil.
Also, the loading substance can comprise an algal oil.

[0071] Suitable polymer components for the disclosed emulsions can be any
of those disclosed herein that are not animal by-products. Many examples
of these are mentioned elsewhere herein.

Method of Making Microcapsules

[0072] Several variables affect the processes of preparing microcapsules
in general, for example, the type of shell material, charge density,
concentration, the ratio of various shell materials, a shell material's
molecular weight (Mw) and distribution, the pH and temperature of the
system, and microion concentration. In the methods disclosed herein, a
non-animal by-product is used as a shell material(s). Many suitable
non-animal by-products are disclosed herein, and they often behave
differently when used to prepare microcapsules as compared to animal
derived shell materials. For example, most vegetable proteins are
globular and are different that animal derived gelatins in terms of
molecular weight, structure, amino acid composition, charge density, and
the like. Gelatin is a protein that can form thermo-reversible gels
through the formation of hydrogen-bond-stabilized triple helices as the
gelatin solution is cooled. Vegetable proteins, like soy proteins, are
more rigid in structure, more heat-stable compared to gelatin, and
denature under prolonged heating, especially above 85° C. Their
amino acid compositions are different, too. See e.g., Table 4.

So, soy proteins require different pH, temperature, concentration, ratio
of polyelectrolytes and microion concentration than gelatin for forming
microcapsules via complex coacervation. Also, because soy proteins
contain more glutamate and lysine residues than gelatin, they are
potentially more active than gelatin for the cross-linking reaction by
transglutaminase, which catalyzes the acyl transfer reaction between
glutaminyl residues and primary amines. Thus, vegetable proteins
microcapsules can be thermal crosslinked by heating up to about
80° C. Similar considerations also apply when using whey or pea
proteins, agar, alginates, gellan gum, gum arabic, xanthan gum, cesains,
and other shell materials that are disclosed herein that are not animal
by-products.

[0073] Since vegetable proteins are not typically cold setting gelling
agents, vegetarian gelatin substitutes, such as pectin, agar, gellan gum,
gum arabic, and alginate, can be used as anionic polysaccharides to
prepare vegetarian microcapsule shells through complex coacervation with
soy proteins. Caseinates or other anionic proteins can also be used
instead of anionic polysaccharides. Again, these vegetarian gelatin
substitutes are different to polyanions use for preparing gelatin
microcapsules in terms of charge density, molecular weight and molecular
weight distribution. Consequently, they require different concentration,
microion concentration, pH, and temperature during complex coacervation
with vegetable proteins.

[0074] Microcapsules prepared by the processes disclosed herein typically
have a combination of payload and structural strength that are suitable
for food articles, supplements, formulation vehicles, and methods
disclosed herein. In one example, the methods disclosed in U.S. Pat. Nos.
6,974,592 and 6,969,530, and US Publication No. 2005-0019416-A1, which
are incorporated by reference in their entirety, can be used to prepare
microcapsules. It is also contemplated that one or more additional shell
layers can be placed on the outer shell of the single-core or multicore
microcapsules. In one example, the techniques described in International
Publication No. WO 2004/041251 A1, which is incorporated by reference in
its entirety, can be used to add additional shell layers to the
single-core and multi-core microcapsules.

[0075] In general, suitable microcapsules can be prepared by a process
that comprises providing an emulsion comprising a first polymer component
a loading substance, and a second polymer component, wherein the first
and second polymer components do not comprise animal by-products;
adjusting pH, temperature, concentration, mixing speed, or a combination
thereof to form an aqueous mixture comprising a primary shell material,
wherein the primary shell material comprises the first and second polymer
components and surrounds the loading substance; cooling the aqueous
mixture to a temperature above the gel point of the primary shell
material until the primary shell material forms agglomerations; and
further cooling the aqueous mixture to form an outer shell around the
agglomeration. In a further example, the agglomeration can be contacted
with a third polymer component; adjusting the pH, temperature,
concentration, mixing speed, or a combination thereof to form an
additional outer shell around the agglomeration. This process can be a
two step process, i.e., the first polymer component and loading substance
can be emulsified and then the second polymer component can be added.
Alternatively, this process can be a one step process, i.e., the first
and second polymer components and the loading substance can be emulsified
together.

[0076] In these methods, the first polymer component, second polymer
component, and third polymer component can be the same as any of the
primary and outer shell materials described herein. That is, the first,
second, and third polymer components can become the primary and/or outer
shell materials in the disclosed methods for preparing microcapsules.
Furthermore, any of the loading substances described herein can be used
in these methods for preparing microcapsules.

[0077] In the disclosed methods, an aqueous mixture of a loading
substance, a first polymer component of the shell material, and a second
polymer component of the shell material is formed. The aqueous mixture
can be a mechanical mixture, a suspension, or an emulsion. When a liquid
loading substance is used, particularly a hydrophobic liquid, the aqueous
mixture can be an emulsion of the loading substance and the polymer
components. In another example, a first polymer component is provided in
aqueous solution, optionally with processing aids, such as antioxidants.
A loading substance can then be dispersed into the aqueous mixture, for
example, by using a homogenizer. If the loading substance is a
hydrophobic liquid, an emulsion is formed in which a fraction of the
first polymer component begins to deposit around individual droplets of
loading substance to begin the formation of primary shells. If the
loading substance is a solid particle, a suspension is formed in which a
fraction of the first polymer component begins to deposit around
individual particles to begin the formation of primary shells. At this
point, another aqueous solution of a second polymer component can be
added to the aqueous mixture (or alternatively, the aqueous mixture can
be added to the aqueous solution of the second polymer component).

[0078] In the processes for preparing microcapsules disclosed herein,
providing an emulsion of the first polymer component and the loading
substance can be accomplished by methods and apparatus known in the art,
e.g., homogenization and high pressure/high shear pumps. For example,
emulsification can take place by emulsifying at from about 1,000 to about
15,000 rpm. The emulsification step can be monitored by removing a sample
of the mixture and analyzing it under such methods as microscopy, light
scattering, turbidity, etc. Generally, emulsification can be performed
until an average droplet size of less than about 1,000, 750, 500, 100, or
10 nm is obtained. Not wishing to be bound by theory but it is believed
that by varying the emulsification speed it is possible to produce single
or multi-core microcapsules. For example, when lower emulsification
speeds are used (e.g., 1,000 to 2,000 rpm), the droplets of the loading
substance are large enough to form a single particle, which upon
encapsulation, produces a single core microcapsule. Conversely, if high
emulsification speeds are used (e.g., 5,000 to 15,000 rpm), the resultant
droplets of loading substance are usually small (e.g., from 1 to 10
μm). These tiny droplets can have higher surface energy and can
readily form agglomerations when pH and/or temperature is adjusted
accordingly, which results in the formation of multi-core microcapsules
upon encapsulation. Particle size can be measured using any typical
equipment known in the art, for example, a COULTER® LS230 Particle
Size Analyzer, Miami, Fla. USA.

[0079] The emulsification step can be performed at less than or greater
than room temperature, e.g., at 4, 10, 15, 20, 30, 37, 40, 50, 60, 70, or
80° C., where any of the stated values can form an upper or lower
endpoint when appropriate. Specific examples include emulsifying the
mixture at from about 10° C. to about 60° C. or from about
30° C. to about 50° C.

[0080] It is further contemplated that antioxidants and/or surfactants,
which are also described herein, can be added to the emulsion and/or
aqueous mixture. Such antioxidants and/or surfactants can be added
before, during, and/or after the emulsion is provided. Further, in the
whole system involving the loading substance, shell materials,
antioxidants, and additional compositions, the antioxidative capacity is
at a certain level when the amount of antioxidants used is given.
Therefore, in the methods for preparing microcapsules disclosed herein,
purging with inert gas such as nitrogen during any or all of
emulsification, mixing, coacervation, and or cooling processes can
prevent the consumption of antioxidants by oxygen from air, and delay
oxidation of the loading substance during storage. It can also prevent
the formation of off-flavor compounds due to oxidation in the
microencapsulation process.

[0081] Also contemplated is that chelators can be added to the emulsion
and/or aqueous mixture. Autoxidation of lipids is catalyzed by metal
ions, particularly iron and copper ions. Thus, chelating of the metal
ions can help retard the oxidation and extend its "lag phase," therefore
extending the shelf-life of bulk oil or encapsulated oils. Like
antioxidants, the chelators can be added before, during and/or after the
emulsion is provided. Examples of suitable chelators include, but are not
limited to are disodium ethylenediamine tetraacetic acid, which is one of
the most frequently used chelating agents in food processing, citric
acid, phytic acid, malic acid, tartaric acid, oxalic acid, succinic acid,
polyphosphoric acids etc.

[0082] The amount of the first and second polymer components of the shell
material provided in the aqueous mixture is typically sufficient to form
both the primary shells and the outer shells of the loading agglomeration
of microcapsules. The loading substance can be provided in an amount of
from about 1% to about 15% by weight of the aqueous mixture, from about
3% to about 8% by weight, or about 6% by weight.

[0083] The pH, temperature, concentration, mixing speed, or a combination
thereof can be adjusted to form an aqueous mixture comprising a primary
shell material, wherein the primary shell material comprises the first
and second polymer components and surrounds the loading substance. If
there is more than one type of polymer component (i.e., the first and
second polymer components are different polymers), complex coacervation
will occur between the components to form a coacervate, which further
deposits around the loading substance to form primary shells of shell
material. The pH adjustment depends on the type of shell material to be
formed. For example, the pH may be adjusted to a value from about 3.5 to
about 5.0, or from about 4.0 to about 5.0. If the pH of the mixture
starts in the desired range, then little or no pH adjustment is required.
In one example, the pH is adjusted to from about 3.5 to about 4.1, from
about 3.6 to about 4.0, or from about 3.7 to about 3.9.

[0084] The initial temperature of the aqueous mixture can be from about
4° C. to about 60° C., or about 10° C. to about
50° C.

[0085] Mixing can be adjusted so that there is good mixing without
breaking the microcapsules as they form. Particular mixing parameters
depend on the type of equipment being used. Any of a variety of types of
mixing equipment known in the art may be used. In one example, an axial
flow impeller, such as LIGHTNIN® A310 or A510, can be used.

[0086] In many examples disclosed herein, the primary shell and the outer
shell of the disclosed microcapsules can comprise a complex coacervate.
The complex coacervate can be formed from the first and second polymer
components. For example, the primary shell and the outer shell can
comprise a complex coacervate between whey protein isolate and agar. All
combinations of first and second polymer components are contemplated
herein for the complex coacervate and the primary and outer shell.

[0087] The aqueous mixture can then be cooled under controlled cooling
rate and mixing parameters to permit agglomeration of the primary shells
to form encapsulated agglomerations of primary shells. Not wishing to be
bound by theory, the encapsulated agglomerations are discrete particles
themselves. It is advantageous to control the formation of the
encapsulated agglomerations at a temperature above the gel point of the
shell material, and to let excess shell material form a thicker outer
shell. It is also possible at this stage to add more polymer (e.g., a
third polymer component), where the polymer is the same or different as
the shell material being used, in order to thicken the outer shell and/or
produce microcapsules having primary and outer shells of different
composition. The outer shell encapsulates the agglomeration of primary
shells to form a rigid encapsulated agglomeration of microcapsules.

[0088] Cooling the aqueous mixture can be accomplished by methods known in
the art (e.g., the use of a chiller). The rate of cooling can be about
1° C. per about 1 to about 100 minutes. For example, the rate of
cooling can be about 1° C. per about 1, 5, 10, 15, 20, 25, 30, 35,
40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 minutes, where any
of the stated values can form an upper or lower endpoint when
appropriate. In specific examples the rate of cooling can be about
1° C./5 minutes. Cooling can take place until the mixture reaches
a temperature of from about 5° C. to about 10° C., e.g.,
about 5° C.

[0089] Processing aids can be included in the shell material (e.g.,
primary and/or outer shells). Processing aids can be used for a variety
of reasons. For example, they may be used to promote agglomeration of the
primary microcapsules, stabilize the emulsion system, improve the
properties of the outer shells, control microcapsule size, and/or to act
as an antioxidant. In one aspect, the processing aid can be an
emulsifier, a fatty acid, a lipid, a wax, a microbial cell (e.g., yeast
cell lines), a clay, or an inorganic compound (e.g., calcium carbonate).
Not wishing to be bound by theory, these processing aids can improve the
barrier properties of the microcapsules. In one aspect, one or more
antioxidants can be added to the shell material. Antioxidant properties
are useful both during the process (e.g., during coacervation and/or
spray drying) and in the microcapsules after they are formed (i.e., to
extend shelf-life, etc). Preferably a small number of processing aids
that perform a large number of functions can be used. In one aspect, the
antioxidant can be a phenolic compound, a plant extract, or a
sulfur-containing amino acid. In one aspect, ascorbic acid or citric acid
(or a salt thereof such as sodium or potassium ascorbate or sodium or
potassium citrate) can be used to promote agglomeration of the primary
microcapsules, to control microcapsule size and to act as an antioxidant.
The antioxidant can be used in an amount of about 100 ppm to about 12,000
ppm, or from about 1,000 ppm to about 5,000 ppm. Other processing aids
such as, for example, metal chelators, can be used as well. For example,
ethylene diamine tetraacetic acid can be used to bind metal ions, which
can reduce the catalytic oxidation of the loading substance.

[0090] In the disclosed microcapsules, the shell material can also be
cross-linked. Thus, the disclosed methods can further involve the
addition of a cross-linker. The cross-linker can be added to further
increase the rigidity of the microcapsules by cross-linking the shell
material in both the outer and primary shells and to make the shells
insoluble in both aqueous and oily media. In one example, the
cross-linker is added after the outer shell of the microcapsule is
produced. Any suitable cross-linker can be used and the choice of
cross-linker can vary depending upon the selection of the first and
second polymer component. In another example, the cross-linkers can be
enzymatic cross-linkers (e.g. transglutaminase), aldehydes (e.g.
formaldehyde or glutaraldehyde), tannic acid, alum or a mixture thereof.
In another aspect, the cross-linker can be a plant extract or a phenolic.
It is also contemplated that one or more loading substances (e.g.,
antioxidants) can be used with the cross-linker. When the microcapsules
are to be used in a formulation that is to be delivered to an organism,
the cross-linkers are preferably non-toxic or of sufficiently low
toxicity. The amount of cross-linker used depends on the components
selected and can be adjusted to provide more or less structural rigidity
as desired. In one aspect, the amount of cross-linker that can be used is
in the amount of about 0.1% to about 5.0%, about 0.5% to about 5.0%,
about 1.0% to about 5.0%, about 2.0% to about 4.0%, or about 2.5%, by
weight of the first polymer component. In general, one skilled in the art
can routinely determine the desired amount in any given case by simple
experimentation. The cross-linker can be added at any stage of the
process; however, it can typically be added after the cooling step.

[0091] Further, in some applications, the use of transglutaminase to
crosslink the microcapsules may not be desired (e.g., the temperature and
pH are too low and/or the transglutaminase is expensive). Thus, it is
contemplated herein that the use of glutaraldehyde can be in the
disclosed methods to cross-link the disclosed microcapsules. In certain
examples, the use of one or more compositions comprising an amino acid or
protein, can react with residual glutaraldehyde that was totally or
partially unreacted from the crosslinking reaction. That is, unreacted
and half reacted glutaraldehyde (i.e., with one aldehyde group still
reactive) can be neutralized by the ε-amino group of lysine or
other amino groups on proteins, making the final product safer. In this
sense, the compositions comprising amino acids and/or proteins can
improve the microcapsule shell by filling any pores and neutralize
glutaraldehyde from the crosslinking reaction. This approach can also
eliminate the need to wash the microcapsule after crosslinking since the
microcapsule will be essentially free of glutaraldehyde. Crosslinking can
also be accomplished with genipin (e.g., with genipin and carboxylmethyl
chitosan).

[0092] It is also possible to crosslink the disclosed microcapsules with
heat. For example, heating to about 80° C. for 30 minutes or
heating to 95° C. for 5 minutes can effectively crosslink the
disclosed microcapsules.

[0093] Further, the disclosed microcapsules can be washed with water
and/or dried to provide a free-flowing powder. Thus, the disclosed
methods of preparing microcapsules can comprise a drying step for the
microcapsules. Drying can be accomplished by a number of methods known in
the art such as, for example, freeze drying, drying with ethanol, or
spray drying. In one aspect, spray drying can be used for drying the
microcapsules. Spray drying techniques are disclosed in "Spray Drying
Handbook", K. Masters, 5th edition, Longman Scientific Technical UK,
1991, the disclosure of which is hereby incorporated by reference at
least for its teaching of spray drying methods.

[0095] Drying agents or anticaking agents can be used to help produce free
flowing powders. Typically, drying agents have high porosity, which can
help adsorb surface oil and flavor compounds due to the raw materials, or
the oxidation of lipids. Examples of suitable drying and/or anticaking
agents include, but are not limited to, HUBERSORB® and ZEOTHIX® (J.
M. Huber Corp; Harve de Grace, Md.) and CAPSUL® (from National Starch
& Chemical Co.) and VITACEL® (J. Rettenmair USA; Schoolcraft, Mich.).

[0096] Incorporating Antioxidants into the Powder

[0097] In other examples, disclosed herein are methods for incorporating
antioxidants into and/or onto the primary shell, the outer shell(s), or
both primary and outer shell(s). materials. The disclosed methods
comprise providing a microcapsule as disclosed herein, providing an
emulsion comprising a polymer component and an antioxidant; combining the
emulsion and the microcapsule, to thereby provide a microcapsule with a
shell material comprising the antioxidant. The resulting suspension can
then be cooled and the coated microcapsules can be dried. In many
suitable examples, the microcapsules can be included in a slurry that
contains the antioxidants and the slurry can be spray dried. Suitable
antioxidants include, but are not limited to, CoQ10, lutein, zeaxanthan,
carotene, and combinations thereof. These can be used alone or in
addition to the amino acids, proteins, saccharides, or waxes disclosed
herein.

Formulation Vehicles

[0098] Also disclosed herein are formulation vehicles comprising the
microcapsules disclosed herein. Any of the microcapsules described herein
can be incorporated into a formulation vehicle. Examples of formulation
vehicles are provided herein and include, but are not limited to,
foodstuffs, beverages, nutraceutical formulations, pharmaceutical
formulations, lotions, creams, or sprays. In some other specific
examples, the disclosed emulsions and/or microcapsules can be
incorporated into gels, gel capsules, or tablets. Other vehicles include
powders or powders coated with a polymer. Such vehicles can be given
orally or, in the case of powders for example, sprinkled onto food or
beverages.

[0099] Supplements

[0100] Also, disclosed herein are nutritional supplements that comprise
the microcapsules disclosed herein. A nutritional supplement is any
compound or composition that can be administered to or taken by a subject
to provide, supply, or increase a nutrient(s) (e.g., vitamin, mineral,
essential trace element, amino acid, peptide, nucleic acid,
oligonucleotide, lipid, cholesterol, steroid, carbohydrate, and the
like). For example, a nutritional supplement can comprise a composition
comprising one or more loading substances disclosed herein.

[0101] The nutritional supplement can comprise any amount of the
microcapsules disclosed herein, but will typically contain an amount
determined to supply a subject with a desired dose of a loading substance
(e.g., EPA and/or DHA). The exact amount of microcapsules required in the
nutritional supplement will vary from subject to subject, depending on
the species, age, weight and general condition of the subject, the
severity of any dietary deficiency being treated, the particular mode of
administration, and the like. Thus, it is not possible to specify an
exact amount for every nutritional supplement. However, an appropriate
amount can be determined by one of ordinary skill in the art using only
routine experimentation given the teachings herein.

[0102] The nutritional supplement can also comprise other nutrient(s) such
as vitamins other trace elements, minerals, and the like. Further, the
nutritional supplement can comprise other components such as
preservatives, antimicrobials, anti-oxidants, chelating agents,
thickeners, flavorings, diluents, emulsifiers, dispersing aids, or
binders.

[0103] The nutritional supplements are generally taken orally and can be
in any form suitable for oral administration. For example, a nutritional
supplement can typically be in a tablet, gel-cap, capsule, liquid,
sachets, or syrup form.

[0104] The nutritional supplements can be designed for humans or animals,
based on the recommended dietary intake for a given individual. Such
considerations are generally based on various factors such as species,
age, and sex as described above, which are known or can be determined by
one of skill in the art. In one example, the disclosed supplements can be
used as a component of feed for animals such as, but not limited to,
livestock (e.g., pigs, chickens, cows, goats, horses, and the like) and
domestic pets (e.g., cats, dogs, birds, and the like).

[0105] Pharmaceutical Formulations

[0106] Also, pharmaceutical formulations comprising the disclosed
microcapsules are disclosed herein. A suitable pharmaceutical formulation
can comprise any of the disclosed compositions with a pharmaceutically
acceptable carrier. For example, a pharmaceutical formulation can
comprise one or more of the disclosed microcapsules and a
pharmaceutically acceptable carrier. The disclosed pharmaceutical
formulations can be used therapeutically or prophylactically.

[0107] By "pharmaceutically acceptable" is meant a material that is not
biologically or otherwise undesirable, i.e., the material can be
administered to a subject without causing any undesirable biological
effects or interacting in a deleterious manner with any of the other
components of the pharmaceutical formulation in which it is contained.
The carrier would naturally be selected to minimize any degradation of
the active ingredient and to minimize any adverse side effects in the
subject, as would be well known to one of skill in the art.

[0108] Pharmaceutical carriers are known to those skilled in the art.
These most typically would be standard carriers for administration of
drugs to humans, including solutions such as sterile water, saline, and
buffered solutions at physiological pH. Suitable carriers and their
formulations are described in Remington: The Science and Practice of
Pharmacy, 21st ed., Lippincott Williams & Wilkins, Philadelphia,
Pa., 2005, which is incorporated by reference herein for its teachings of
carriers and pharmaceutical formulations. Typically, an appropriate
amount of a pharmaceutically-acceptable salt is used in the formulation
to render the formulation isotonic. Examples of the
pharmaceutically-acceptable carrier include, but are not limited to,
saline, Ringer's solution and dextrose solution. The pH of the solution
can be from about 5 to about 8 (e.g., from about 7 to about 7.5). Further
carriers include sustained release preparations such as semipermeable
matrices of solid hydrophobic polymers containing the disclosed
compounds, which matrices are in the form of shaped articles, e.g.,
films, liposomes, microparticles, or microcapsules. It will be apparent
to those persons skilled in the art that certain carriers can be more
preferable depending upon, for instance, the route of administration and
concentration of composition being administered. Other compounds can be
administered according to standard procedures used by those skilled in
the art.

[0109] Pharmaceutical formulations can include additional carriers, as
well as thickeners, diluents, buffers, preservatives, surface active
agents and the like in addition to the compounds disclosed herein.
Pharmaceutical formulations can also include one or more additional
active ingredients such as antimicrobial agents, anti-inflammatory
agents, anesthetics, and the like.

[0110] The pharmaceutical formulation can be administered in a number of
ways depending on whether local or systemic treatment is desired, and on
the area to be treated. Administration can be topically (including
ophthalmically, vaginally, rectally, intranasally), orally, by
inhalation, or parenterally, for example by intravenous drip,
subcutaneous, intraperitoneal or intramuscular injection. The disclosed
compounds can be administered intravenously, intraperitoneally,
intramuscularly, subcutaneously, intracavity, or transdermally.

[0111] Preparations for parenteral administration include sterile aqueous
or non-aqueous solutions, suspensions, and emulsions. Examples of
non-aqueous solvents are propylene glycol, polyethylene glycol, vegetable
oils such as olive oil, marine oils, and injectable organic esters such
as ethyl oleate. Aqueous carriers include water, alcoholic/aqueous
solutions, and emulsions or suspensions, including saline and buffered
media. Parenteral vehicles include sodium chloride solution, Ringer's
dextrose, dextrose and sodium chloride, lactated Ringer's, and fixed
oils. Intravenous vehicles include fluid and nutrient replenishers,
electrolyte replenishers (such as those based on Ringer's dextrose), and
the like. Preservatives and other additives may also be present such as,
for example, antimicrobials, anti-oxidants, chelating agents, and inert
gases and the like.

[0112] Pharmaceutical formulations for topical administration may include
ointments, lotions, creams, gels, drops, suppositories, sprays, liquids
and powders. Conventional pharmaceutical carriers, aqueous, powder or
oily bases, thickeners and the like can be desirable.

[0113] Pharmaceutical formulations for oral administration include, but
are not limited to, powders or granules, suspensions or solutions in
water or non-aqueous media, capsules, sachets, or tablets. Thickeners,
flavorings, diluents, emulsifiers, dispersing aids, or binders can be
desirable.

[0116] Also disclosed herein are foodstuffs that comprise any of the
disclosed microcapsules. By "foodstuff" is meant any article that can be
consumed (e.g., eaten, drank, or ingested) by a subject. In one example,
the disclosed compositions can be used as nutritional supplements that
are added to a foodstuff. For example, the disclosed microcapsules can be
added to food or beverages. In this sense, the disclosed compositions can
be prepared in, for example, a powdered form and contained in articles
such as sachets or shakers, which can be used to pour or sprinkle the
disclosed compositions onto and into food and beverages.

[0117] In some examples, the foodstuff is a baked good, a pasta, a meat
product, a frozen dairy product, a milk product, a cheese product, an egg
product, a condiment, a soup mix, a snack food, a nut product, a plant
protein product, a hard candy, a soft candy, a poultry product, a
processed fruit juice, a granulated sugar (e.g., white or brown), a
sauce, a gravy, a syrup, a nutritional bar, a beverage, a dry beverage
powder, a jam or jelly, a fish product, or pet companion food. In other
examples, the foodstuff is bread, tortillas, cereal, sausage, chicken,
ice cream, yogurt, milk, salad dressing, rice bran, fruit juice, a dry
beverage powder, liquid beverage, rolls, cookies, crackers, fruit pies,
or cakes.

Methods of Use

[0118] The disclosed microcapsules also have a wide variety of uses. For
example, disclosed herein are methods of delivering a loading substance
to a subject by administering to the subject a microcapsule as disclosed
herein. Also disclosed is the use a microcapsule as disclosed herein to
prepare a medicament for delivering a loading substance to a subject. The
disclosed microcapsules can be particularly useful for delivering
substances to those on vegan, lactovegetarian, ovo-lactovegetarian,
and/or semi-vegetarian diets.

[0119] The use of microcapsules can protect certain compositions from
oxidation and degradation, keeping the loading substance fresh. Also,
because microcapsules can hide the unpleasant odor or taste of certain
compositions, the methods disclosed herein can be particularly useful for
delivering and supplementing unpleasant compositions. Still further, the
use of microcapsules can allow various loading substances to be added to
food articles which are otherwise not amenable to supplementation. For
example, omega-3 fatty acids can degrade or oxidize in air and can be
sensitive to food preparation techniques (e.g., baking). By the use of
microencapsulated omega-3 fatty acids, these compositions can be added to
food without significant degradation during food preparation.

[0120] Particularly suitable microcapsules include those that are
resistant to rupture during the preparation of the food article
(including packaging, transportation, and storage of the food article).
In some examples, the microcapsules can be of a size and consistency that
does not detract from the texture and constitution of the food article.

[0121] In a particular example, the disclosed microcapsules (including
nutritional supplements, pharmaceutical formulations, delivery devices,
and foodstuffs that contain the disclosed microcapsules) can be used as a
source of fatty acids (e.g., omega-3 fatty acids), lowering triglycerides
and influencing diabetes related biochemistry. In another particular
example, disclosed herein are methods of supplementing omega-3 fatty
acids in a subject by administering an effective amount of a microcapsule
disclosed herein, wherein the loading substance comprises an omega-3
fatty acid. In another example, disclosed herein are methods of lowering
cholesterol levels, triglyceride levels, or a combination thereof in a
subject by administering an effective amount of an emulsion and/or
microcapsule disclosed herein.

[0124] Despite the strong evidence for the benefit of omega-3 fatty acids
like EPA and DHA in prevention of cardiovascular disease, the average
daily consumption of these fatty acids by North Americans is estimated to
be between 0.1 to 0.2 grams, compared to a suggested daily intake of 0.65
grams to confer benefit (Webb, "Alternative sources of omega-3 fatty
acids." Natural Foods Merchandiser 2005, XXVI(8):40-4). Since altering
dietary patterns of populations is difficult and many people do not like
to eat fish, dietary supplementation with EPA and DHA is an important
approach to addressing this problem. Unfortunately, many supplements of
omega-3 fatty acids are sensitive to oxidation and can be foul smelling
and tasting. Further, compliance with dietary supplement regimens
requires discipline, which is often wanting. In light of the health
benefits of omega-3 fatty acids, the disclosed microcapsules can be used
to deliver omega-3 fatty acids to a subject.

[0125] In the disclosed methods of use, the emulsions and/or microcapsules
that are administered can be any of the compositions disclosed herein.
For example, the disclosed microcapsules can be used in the disclosed
methods in the form of any of the nutritional supplements disclosed
herein. In another example, the disclosed microcapsules can be used in
the disclosed methods in the form of any of the pharmaceutical
formulations disclosed herein. In still another example, the disclosed
microcapsules can be incorporated in any of the delivery devices
disclosed herein, or incorporated into any foodstuff disclosed herein and
used in the disclosed methods.

[0126] It is contemplated that the methods disclosed herein can be
accomplished by administering various forms of the disclosed
microcapsules. For example, one can administer any of the pharmaceutical
formulations with any of the foodstuffs disclosed herein. In another
example, one can administer a tablet or capsule with any of the
nutritional supplements disclosed herein. In yet another example, one can
administer any of the pharmaceutical formulations with any of the
delivery devices and nutritional supplement disclosed herein, and the
like.

[0127] Dosage

[0128] When used in the above described methods or other treatments, or in
the nutritional supplements, pharmaceutical formulations, delivery
devices, or foodstuffs disclosed herein, an "effective amount" of one of
the disclosed microcapsules can be employed in pure form or, where such
forms exist, in pharmaceutically acceptable salt form, and with or
without a pharmaceutically acceptable excipient, carrier, or other
additive.

[0129] The specific effective dose level for any particular subject will
depend upon a variety of factors including the disorder being treated and
the severity of the disorder; the identity and activity of the specific
composition employed; the age, body weight, general health, sex and diet
of the patient; the time of administration; the route of administration;
the rate of excretion of the specific composition employed; the duration
of the treatment; drugs used in combination or coincidental with the
specific composition employed and like factors well known in the medical
arts. For example, it is well within the skill of the art to start doses
of a composition at levels lower than those required to achieve the
desired therapeutic effect and to gradually increase the dosage until the
desired effect is achieved. If desired, the effective daily dose can be
divided into multiple doses for purposes of administration. Consequently,
single dose compositions can contain such amounts or submultiples thereof
to make up the daily dose.

[0130] The dosage can be adjusted by the individual physician or the
subject in the event of any counterindications. Dosage can vary, and can
be administered in one or more dose administrations daily, for one or
several days. Guidance can be found in the literature for appropriate
dosages for given classes of pharmaceutical products.

[0131] Further, disclosed are methods for delivering a disclosed
composition to a subject by administering to the subject any of the
nutritional supplements, pharmaceutical formulations, delivery devices,
and/or foodstuffs disclosed herein. The disclosed compositions (including
nutritional supplements, delivery devices, and pharmaceutical
formulations) can typically be administered orally.

EXAMPLES

[0132] The following examples are set forth below to illustrate the
methods and results according to the disclosed subject matter. These
examples are not intended to be inclusive of all aspects of the subject
matter disclosed herein, but rather to illustrate representative methods
and results. These examples are not intended to exclude equivalents and
variations of the present invention which are apparent to one skilled in
the art.

[0133] Efforts have been made to ensure accuracy with respect to numbers
(e.g., amounts, temperature, pH, etc.) but some errors and deviations
should be accounted for. Unless indicated otherwise, parts are parts by
weight, temperature is in ° C. or is at ambient temperature, and
pressure is at or near atmospheric. There are numerous variations and
combinations of conditions, e.g., component concentrations, temperatures,
pressures, and other reaction ranges and conditions that can be used to
optimize the product purity and yield obtained from the described
process. Only reasonable and routine experimentation will be required to
optimize such process conditions.

[0134] Certain materials, compounds, compositions, and components
disclosed herein can be obtained commercially or readily synthesized
using techniques generally known to those of skill in the art. For
example, the starting materials and reagents used in preparing the
disclosed compositions are either available from commercial suppliers
such as Ocean Nutrition Canada, Ltd. (Dartmouth, NS, Canada), Aldrich
Chemical Co., (Milwaukee, Wis.), Acros Organics (Morris Plains, N.J.),
Fisher Scientific (Pittsburgh, Pa.), or Sigma (St. Louis, Mo.) or are
prepared by methods known to those skilled in the art following
procedures set forth in references such as Fieser and Fieser's Reagents
for Organic Synthesis, Volumes 1-17 (John Wiley and Sons, 1991); Rodd's
Chemistry of Carbon Compounds, Volumes 1-5 and Supplements (Elsevier
Science Publishers, 1989); Organic Reactions, Volumes 1-40 (John Wiley
and Sons, 1991); March's Advanced Organic Chemistry, (John Wiley and
Sons, 4th Edition); and Larock's Comprehensive Organic Transformations
(VCH Publishers Inc., 1989).

Example 1

Microencapsulation Using WPI/Agar-WPI/Gellan Gum

[0135] 4.0 g of agar (TIC pretested agar; TIC Gums; Belcamp, Md.) was
added to 100.0 g of boiling water to be hydrated and dissolved. The
resulting solution was then transferred into a 2-L reactor with 600.0 g
of deionized water maintained at 65° C. Next, 1.0 g of sodium
ascorbate was added to the solution in the reactor and the pH was
determined to be about 6.6.

[0136] 35.0 g of WPI (whey protein isolate) (Alacen® 895, NZMP (USA)
Inc., Lemoyne, Pa.) was added to 90.0 g of deionized water under
agitation at room temperature (25° C.). The dissolved WPI was then
cooled to 10° C. 70.0 g of fish oil (XODHA from Ocean Nutrition
Canada, Ltd.; Dartmouth, NS) was added to the cold WPI solution and the
resulting mixture was emulsified by a POLYTRON PT 6100® homogenizer
(Kinematica AG, Lucerne, Switzerland) at 8000 rpm for 5 minutes while the
temperature was maintained at 10° C. The resulting emulsion was
examined under a microscope after emulsification to verify that the oil
droplets were small and uniform (about 1-5 μm in diameter).

[0137] The emulsion was added to the agar solution in the reactor. The pH
value of the resulting mixture was about 6.4. Then, pH was adjusted to
about 5.0 with 10% w/w phosphoric acid to form about 30 μM
agglomerations of primary microcapsules.

[0138] 4.0 g of low acyl gellan gum (Kelcogel F, from CPKELCO; San Diego,
Calif.) and 4.0 g of WPI were dissolved in 600.0 g of deionized water at
about 60° C. The solution pH, which was initially 6.2, was
adjusted to 5.0 with 10% w/w phosphoric acid. This mixture was added to
the microcapsules in the reactor. 3.0 g of CaCl2 in 20.0 g of
distilled water solution was prepared and also added to the suspension of
microcapsules. The resulting slurry was quickly cooled to 20° C.
and agitation speed was increased during cooling to avoid gelling. The
finished suspension of microcapsules was ready for spray drying to
produce a free flowing powder. Such a microcapsule would be suitable for
a lactovegetarian, ovo-lactovegetarian, and semi-vegetarian diet.

Example 2

Microencapsulation Using Whey Protein and Low Methoxyl Pectin

[0139] 29.3 g of whey protein isolate (WPI, Alacen 895, NZMP (USA) Inc.)
was dissolved in 322 g of water in a 2-L reactor with agitation. The
resulting solution was kept at 30° C. while 7 g of sodium
ascorbate was then added. 15 g of fish oil (XO30TG, Ocean Nutrition
Canada, Ltd.) was next added to the WPI solution. The solution was then
emulsified with a POLYTRON PT 6100® homogenizer at 10,000 rpm for 5
minutes. Next, 972 g of distilled water was added to the resulting
emulsion in the reactor while the temperature was maintained at
30° C. 14.6 g of pretested PECTIN LM32® from TIC Gums (Belcamp,
Md.) were dissolved in 168.2 g of distilled water and then added to the
diluted emulsion in the reactor. Suspension pH was adjusted to 3.1 with
10% phosphoric acid (about 50 mL) to form about 10 μm agglomerations
of primary microcapsules. The mixture was then heated from 30° C.
to 85° C. at an average heating rate of 1.3° C. per minute.
The particle size increased to 30 μm and the mixture was cooled to
room temperature naturally and agitated overnight. The finished
suspension of microcapsules was then ready for coating processes, or
spray dried to produce a free flowing powder. Such a microcapsule would
be suitable for a lactovegetarian, ovo-lactovegetarian, and
semi-vegetarian diet.

Example 3

Microencapsulation Using Gelatin and Low Methoxyl Pectin

[0140] 570 g of deionized water was added to a reactor and heated to about
53° C. 8.0 g of low methoxyl pectin (LM-12 CG® from C.P. Kelco;
San Diego, Calif.) was dissolved in 349 g of water at about 53° C.
40.0 g of fish gelatin (240 Bloom, from LAPI; Tuscany, Italy) was
dissolved in 293 g of water at about 53° C. After the gelatin was
completely dissolved, 6.1 g of sodium ascorbate was added to the gelatin
solution. 72.0 g of DHA oil (XODHA, Ocean Nutrition Canada, Ltd.) was
then added to the gelatin solution and the resulting mixture was
emulsified with a POLYTRON PT 6100 homogenizer at 7500 rpm for 4 minutes.
The emulsion was then added to water in the reactor and pH of the
solution was adjusted to 8.04. The pectin solution was then added to the
reactor and coacervation was commenced with the addition of citric acid
until pH 4.52 and the desired particle size was reached (about 30 μm).
The slurry was cooled at 5° C. per minute to 4° C. Once the
slurry reached 4° C., 2.6 g of transglutaminase (ACTIVA TI,
Ajinomoto Co. Inc., Tokyo, JP) was added and pH was adjusted to 5.04.
Temperature was then raised to 25° C. in 30 minutes and maintained
for crosslinking at 25° C. for 12 hours. The slurry of
microcapsules was then ready for use in food or spray drying to produce a
free flowing powder. Such a microcapsule would be suitable for a
semi-vegetarian diet.

Example 4

Microencapsulation Using Gelatin-Alginate (One-Step Process)

[0141] 44.8 g of fish gelatin (240 Bloom, LAPI) was dissolved in 254 g of
water. This solution was then heated to 40° C.

[0142] 1179 g of distilled water was added to a 2-L reactor and
temperature was maintained at 40° C. An amount of 7.5 g ascorbic
acid was added into the reactor. Next, 30 mL of 10% citric acid were
added to the reactor. The solution pH was 3.3. An amount of 10% NaOH
solution was then added to the reactor to reach a pH of 4.8.

[0143] 72.0 g of fish oil (XO30TG, Ocean Nutrition Canada, Ltd.) was added
to the gelatin solution. The resulting solution was then emulsified with
a POLYTRON PT 6100® homogenizer at 7500 rpm for 4 minutes. The
resulting emulsion was examined under a microscope after emulsification
to verify that the oil droplets were small and uniform (about 1-5 μm
in diameter).

[0144] The emulsion was added to distilled water in the reactor and pH of
the mixture was found be 4.9. NaOH was then added to bring the pH to 5.4.

[0145] 3.2 g of alginate (PROTANAL LFR 5/60® from FMC Biopolymer;
Philadelphia, Pa.) was dissolved in 61 g of distilled water. This
alginate solution was then added to the diluted emulsion in the reactor.
The mixture in the reactor had a pH of 5.5 and the oil droplets were 1-3
μm in diameter. Suspension pH was then lowered with 10% citric acid in
order to form agglomerations of primary microcapsules. After pH was
lowered to 5.0 with the addition of 12 mL of acid, the slurry was cooled
to 4° C. with controlled cooling at 5° C. per minute.

[0146] 3.1 g of transglutaminase dissolved in 10 g of distilled water were
added to the microcapsules at 4° C. Temperature was raised to
25° C. in 30 minutes for crosslinking overnight (12 hours). The
finished suspension of microcapsules was then ready for food processes,
or spray dried to produce a free flowing powder. Such a microcapsule
would be suitable for a semi-vegetarian diet.

Example 5

Microencapsulation Using Gelatin-Alginate (Two-Step Process)

[0147] 22.6 g of fish gelatin (240 Bloom, LAPI) was dissolved in 160 g of
water. 7.6 g of sodium ascorbate was then added and the solution was
heated to 40° C. The solution pH was adjusted to 6.0 by adding 10%
NaOH solution.

[0148] 1.4 g of alginate (PROTANAL LFR 5/60®, FMC Biopolymer) was
dissolved in 44 g of distilled water. This alginate solution was then
added to the gelatin solution.

[0149] 569 g of distilled water was added to a 2-L reactor and the
temperature was maintained at 40° C. 69.0 g of fish oil (XO30TG,
Ocean Nutrition Canada, Ltd.) was added to the gelatin and alginate
solution and then emulsified with a POLYTRON PT 6100® homogenizer at
7500 rpm for 3 minutes. The emulsion was examined under a microscope
after emulsification and verified that the oil droplets were small and
uniform (about 1-5 μm in diameter). The emulsion was added to
distilled water in the reactor and the pH value of the mixture was 5.8.
The pH value was then lowered with 10% citric acid in order to form
agglomerations of primary microcapsules. After pH was lowered to 5.1 with
the addition of 4.5 mL of acid, the slurry was cooled to 37° C.

[0150] A gelatin solution and an alginate solution were prepared for the
second step process as follows. 18.6 g of fish gelatin (LAPI) was
dissolved in 251 g of water with 3.0 g of sodium ascorbate. This solution
was then heated to 37° C. 2.3 g of alginate was dissolved in 384 g
of distilled water. The resulting alginate solution was then heated to
37° C. and mixed with the gelatin solution. The mixture was cloudy
and had a pH of 5.1. A 10% NaOH solution was added to the mixture to
bring the pH up to 5.6. The solution became at least partially
transparent. The mixture was added to the slurry of microcapsules in the
reactor with increased agitation to prevent clumping. The slurry was
cooled at 5° C. per minute to 4° C.

[0151] 3.1 g of transglutaminase dissolved in 10 g of distilled water was
added to the slurry at 4° C. Temperature was then increased to
25° C. for crosslinking overnight (12 hours). The finished
suspension of microcapsules was then ready for food processes, or spray
dried to produce a free flowing powder. Such a microcapsule would be
suitable for a semi-vegetarian diet.

Example 6

Microencapsulation Using SPI/Agar-SPI/Gellan Gum

[0152] 4.0 g of agar was added to 100.0 g of boiling water to be hydrated
thoroughly. The solution was then transferred into a 2-L reactor with
600.0 g of distilled water maintained at 65° C.

[0153] 45.0 g of soy protein isolate (SPI) (ICN Biomedicals, Inc.; Irving,
Calif.) was added to 300.0 g of distilled water under agitation and
warmed to 65° C. to dissolve. 60.0 g of fish oil (XODHA, Ocean
Nutrition Canada, Ltd.) was added to the SPI solution. The resulting
mixture was then emulsified with a POLYTRON PT 6100 homogenizer at 8000
rpm for 8 minutes. The emulsion was examined under a microscope after
emulsification to verify that the oil droplets were about 5 μm in
diameter. The emulsion was then added to the agar solution in the reactor
and the pH of the mixture was about 6.7. The pH was adjusted to 5.0 with
10% w/w phosphoric acid to form about 30 μm agglomerations of primary
microcapsules.

[0154] 4.0 g of low acyl gellan gum and 8.0 g of SPI were dissolved in
400.0 g of distilled water at about 60° C. The pH was adjusted
from 6.6 to 5.0 with 10% w/w phosphoric acid. The resulting mixture was
added to the suspension of microcapsules in the reactor. 1.5 g of
CaCl2 in 10.0 g of distilled water was then added to the reactor.
The resulting slurry was then quickly cooled to 20° C. and
agitation speed was increased during cooling to avoid gelling. The
finished suspension of microcapsules was ready for spray drying to
produce a free flowing powder. Such a microcapsule would be suitable for
a vegan, lactovegetarian, ovo-lactovegetarian, and semi-vegetarian diet.

Example 7

Microencapsulation Using SPI/Agar/Gellan Gum with ARA Oil

[0155] 40.0 g soy protein isolate (ICN Biomedicals, Inc.) was dissolved in
330.0 g of distilled water. The resulting solution was heated up to
60° C. and pH was adjusted to about 11.

[0156] 60.0 g of ARA oil (Wuhan Fuxing Biotechnology Pharmaceutical Co.
Ltd., Wuhan, China) was heated to 50° C. The ARA oil was then
added to the soy protein solution and emulsified at 8000 rpm for 5
minutes. The emulsion was examined under a microscope after
emulsification to verify that the oil droplets were about 1-2 μm in
diameter.

[0157] 3.0 g of agar (TIC pretested agar, TIC Gums; Belcamp, Md.) was
dissolved in 100.0 g of boiling distilled water and then transferred to a
2-L reactor with 600.0 g distilled water and 5.0 g of sodium ascorbate.
Temperature was maintained at 55° C. and the mixture had a pH of
about 7.0.

[0158] The emulsion was then added to the reactor and the pH of the
mixture was about 10.8. The pH value was then adjusted to about 5.7 with
10% phosphoric acid to form about 40 μm agglomerations of the primary
microcapsules.

[0159] 3.2 g of transglutaminase in 10.0 g of distilled water was added to
the reactor and the suspension was maintained at 50° C. for 3
hours before being cooled down to 44° C. 5.0 g of gellan gum
(Kelcogel F, CPKELCO) and 2.0 g of sodium ascorbate were dissolved in
400.0 g of distilled water at 65° C. and then cooled to 50°
C. 4.0 g of SPI was dissolved in 50.0 g of distilled water with pH
adjusted to about 9. The SPI solution was then mixed with the gellan gum
solution and the pH value was adjusted to about 6.7. The resulting
SPI/gellan gum solution was then added to the agglomerated primary
microcapsules in the reactor at 44° C.

[0160] 1.60 g CaCl2 in 10.0 g distilled water was added to the
reactor and agitation speed was increased gradually as the solution was
quickly cooled down to 20° C. The finished suspension of
microcapsules had a compact structure and shell, and the shell survived
after boiling. Such a microcapsule would be suitable for a vegan,
lactovegetarian, ovo-lactovegetarian, and semi-vegetarian diet.

Example 8

Microencapsulation Using SPI/Agar/Gellan Gum with Algal Oil

[0161] 26.67 g of soy protein isolates (ICN Biomedicals, Inc.) was
dissolved in 220.0 g of distilled water. The resulting solution was
heated up to 60° C. and the pH was adjusted to 10.6.

[0162] 40.0 g of algal oil (DHASCO-S from Martek Biosciences; Columbia,
Md.) was heated to 50° C. The algal oil was then added to the soy
protein solution and emulsified at 8000 rpm for 5 minutes. The emulsion
was examined under a microscope after emulsification to verify that the
oil droplets were about 1 μm in diameter.

[0163] 2.0 g of agar (TIC pretested agar, TIC Gums; Belcamp, Md.) was
dissolved in 66.7.0 of boiling distilled water and then transferred to a
2-L reactor with 400.0 g of distilled water and 3.33 g of sodium
ascorbate. The temperature in the reactor was maintained at 55° C.
and the mixture had a pH of about 7.0.

[0164] The algal oil emulsion was added to the distilled water in the
reactor and the pH of the mixture was about 10.2. The pH was then
adjusted to about 5.7 with 10% w/w phosphoric acid to form about 30 μm
agglomerations of the primary microcapsules.

[0165] 2.1 g of transglutaminase in 10.0 g of distilled water was next
added to the reactor and the mixture was maintained at 50° C. for
3 hours before cooling down to 44° C.

[0166] 2.67 g of gellan gum (Kelcogel F) and 1.33 g of sodium ascorbate
were dissolved in 266.7 g of distilled water at 65° C. and then
cooled to 50° C. 2.6 g of SPI was dissolved in 30.0 g of distilled
water with pH adjusted to about 9. The SPI solution was then mixed with
the gellan gum solution and pH was adjusted to about 6.7. The resulting
SPI/gellan gum solution was then added to the agglomerated primary
microcapsules in the reactor at 44° C.

[0167] 1.0 g of CaCl2 in 5.0 g distilled water was added to the
reactor and the agitation speed was gradually increased as the solution
was quickly cooled down to 20° C. The finished suspension of
microcapsules had a compact structure and shell, and the shell survived
after boiling. Such a microcapsule would be suitable for a vegan,
lactovegetarian, ovo-lactovegetarian, and semi-vegetarian diet.

Example 9

Microencapsulation Using SPI/Agar/Gellan Gum with Omega-3 Oil

[0168] 8.9 g of soy protein isolates (ICN Biomedicals, Inc.) was dissolved
in 73.3 g of distilled water. The resulting solution was heated to
60° C. and pH was adjusted to 10.6.

[0169] 13.3 g of omega-3 oil (ONC-T18, Ocean Nutrition Canada Ltd.) was
heated to 70° C. and then added to the soy protein solution and
emulsified at 8000 rpm for 5 minutes. The emulsion was examined under a
microscope after emulsification to verify that the oil droplets were
about 1-2 μm in diameter.

[0170] 0.67 g of agar (TIC pretested agar, TIC Gums; Belcamp, Md.) was
dissolved in 22.2 g of boiling distilled water and transferred to a 500
mL reactor with 133.3 g of distilled water and 1.11 g of sodium
ascorbate. Temperature was maintained at 55° C. and the mixture
had a pH of about 7.0. The omega-3 oil emulsion was then added to the
reactor and pH of the mixture was found to be about 10.8. The pH value
was then adjusted to about 5.7 with 10% phosphoric acid to form about 30
μm agglomerations of the primary microcapsules.

[0171] 0.71 g of transglutaminase in 5.0 g of distilled water was added to
the reactor and the temperature was maintain at 50° C. for 3 hours
before being cooled down to 44° C.

[0172] 0.89 g of gellan gum (Kelcogel F, CPKELCO) and 0.44 g of sodium
ascorbate were dissolved in 89.0 g of distilled water at 65° C.
and then cooled to 50° C. 0.89 g of SPI was dissolved in 10.0 g of
distilled water at apH of about 9. The SPI solution was mixed with the
gellan gum solution and pH was adjusted to about 6.7. The SPI/gellan gum
solution was then added to the agglomerated primary microcapsules in the
reactor at 44° C.

[0173] 0.33 g of CaCl2 in 3.0 g of distilled water was added to the
reactor, and the agitation speed was gradually increased as the mixture
was quickly cooled down to 20° C. The finished suspension of
microcapsules had a compact structure and shell, and the shell did not
change after boiling. Such a microcapsule would be suitable for a vegan,
lactovegetarian, ovo-lactovegetarian, and a semi-vegetarian diet.

[0175] 67.0 g of fish oil (XODHA from Ocean Nutrition Canada, Ltd.;
Dartmouth, NS) was heated to 50° C. to be melted and then added to
the WPI solution. The resulting mixture was cooled to 10° C. and
emulsified by a POLYTRON PT 6100® homogenizer (Kinematica AG, Lucerne,
Switzerland) at 8000 rpm for 5 minutes while the temperature was
maintained at 10° C. The resulting emulsion was examined under a
microscope after emulsification to verify that the oil droplets were
small and uniform (about 1-2 μm in diameter).

[0176] The emulsion was added to a 1.5 L reactor with 1200.0 g distilled
water and 6.7 g sodium ascorbate at room temperature. The pH value of the
resulting mixture was about 6.4. pH was then adjusted to about 3.9 with
10% w/w phosphoric acid to form about 30 μm agglomerations of primary
microcapsules.

[0177] The resulting suspension was heated up to 95° C. and held
for 5 minutes, then cooled to room temperature. The finished suspension
of microcapsules was spray dried to produce a free flowing powder with a
compact structure. The induction period was greater The solution was then
emulsified at 9300 rpm for 4 minutes. The emulsion was examined under a
microscope after emulsification to verify that the oil droplets were
small (around 4 μm in diameter).

[0178] 10.0 g of sodium caseinate (NZMP ALANATE 180) was dissolved in a
1.5 L reactor with 957.0 g of distilled water and 6.3 g of sodium
ascorbate at room temperature. The solution in the reactor had a pH
around 6.4. The PPI and fish oil emulsion was added to the sodium
caseinate solution in the reactor and the pH of the mixture was around
6.4.

[0179] pH was then adjusted to about 5.0 with 20% w/w phosphoric acid to
form about 30 μm agglomerations of the primary microcapsules. The
suspension was heated up to 95° C. and held for 10 minutes, then
cooled down to room temperature.

[0180] The finished suspension of microcapsules with compact structure and
round particles was provided after spray-dry. Such a microcapsule would
be suitable for a lactovegetarian, ovo-lactovegetarian, and
semi-vegetarian diet.

Example 13

Microencapsulation Using WPI-Sodium Caseinate

[0181] 40.0 g WPI (Davisco, Bipro) and 10.0 g of sodium caseinate (NZMP
ALANATE 180) were dissolved in 140.0 g of distilled water at room
temperature. 75.0 g of fish oil (Ocean Nutrition Canada) was heated to
50° C. to be melted and then added to the above mixture. The
resultant mixture was then emulsified at 9300 rpm for 5 minutes at
6° C. The emulsion was examined under a microscope after
emulsification to verify that the oil droplets were small (around 2 μm
in diameter).

[0182] 5.3 g of sodium ascorbate was dissolved in a 1.5 L reactor with
800.0 g of distilled water at room temperature. The solution in the
reactor had a pH around 7.4. Then the WPI-sodium caseinate-fish oil
emulsion was added to the sodium ascorbate solution in the reactor and
the pH of the mixture was around 6.5.

[0183] pH was then adjusted to about 4.7 with 20% w/w phosphoric acid to
form about 30 μM agglomerations of the primary microcapsules. The
suspension was heated to 90° C. and held for 20 minutes, then
cooled down to room temperature.

[0184] The finished suspension of microcapsules with compact structure,
and free flowing powder was provided after spray-dry. Such a microcapsule
would be suitable for a lactovegetarian, ovo-lactovegetarian, and
semi-vegetarian diet.

Example 14

Microencapsulation Using Gelatin-Sodium Caseinate

[0185] 40.0 g pork gelatin (Nitta) and 10.0 g of sodium caseinate (NZMP
ALANATE 180) were dissolved in 293.0 g distilled water at 50° C.
75.0 g of fish oil (Ocean Nutrition Canada) was heated to 50° C.
to be melted and was then added to the gelatin solution. The mixture was
then emulsified at 9000 rpm for 4 minutes. The emulsion was examined
under a microscope after emulsification to verify that the oil droplets
were small (around 1.3 μm in diameter).

[0186] In a 1.5 L reactor with 800.0 g distilled water, 6.3 g sodium
ascorbate was added and the temperature was maintained at 45° C.
The solution in the reactor had a pH around 6.4. The gelatin-fish oil
emulsion was then added to the sodium ascorbate solution in the reactor
and the pH of the mixture was around 5.8.

[0187] pH was then adjusted to about 4.7 with 20% w/w phosphoric acid to
form about 20 μm agglomerations of the primary microcapsules. Complex
coacervates were provided. Such a microcapsule would be suitable for a
lactovegetarian, ovo-lactovegetarian, and semi-vegetarian diet.

[0188] It will be apparent to those skilled in the art that various
modifications and variations can be made in the present invention without
departing from the scope or spirit of the invention. Other embodiments of
the invention will be apparent to those skilled in the art from
consideration of the specification and practice of the invention
disclosed herein. It is intended that the specification and examples be
considered as exemplary only, with a true scope and spirit of the
invention being indicated by the following claims.